DNA vectors having specialized secondary structures for use in treating hypophosphatasia

Abstract

Provided herein are circular, non-integrating, non-viral DNA vectors which express a TNALP polypeptide in an improved expression cassette, wherein the DNA vectors are capable of forming one or more specialized secondary structures, for example an extended cruciform structure, and methods of using such DNA vectors. These DNA vectors having improved expression cassettes provide for increased potency and long-term TNALP transgene expression.

Description

[0001] DNA VECTORS HAVING SPECIALIZED SECONDARY STRUCTURES FOR USE IN TREATING HYPOPHOSPHATASIA CROSS REFERENCE TO RELATED APPLICATIONS This application claims benefit of International Application PCT / US24 / 52858, filed with the United States Receiving Office on October 24, 2024. The entirety of this application is hereby incorporated by reference herein for all purposes. FIELD OF THE DISCLOSURE The present disclosure provides compositions and methods of using circular, double stranded DNA non-viral vectors capable of forming specialized secondary structures, for example extended cruciform structures, for treating hypophosphatasia (HPP). INCORPORATION BY REFERENCE The Sequence Listing XML file named “24012-005WO2_ST26” created on October 24, 2025, and having a size of 191,396 bytes, is hereby incorporated by reference. BACKGROUND OF THE DISCLOSURE Hypophosphatasia (HPP) is a rare inherited systemic metabolic disease caused by mutations in the ALPL gene which encodes the tissue-nonspecific alkaline phosphatase (TNALP) enzyme. Autosomal recessive and dominant forms of the disease result in a range of clinical entities, but HPP is generally characterized by impaired mineralization (“calcification”) of bones and teeth. There are six major clinical forms of HPP—perinatal, infantile, childhood (severe or mild), adult, and odontohypophosphatasia—and severity of the disorder ranges from an extremely severe “perinatal” (at birth) form that can cause stillbirth to a more common (“odonto”) form associated with only early loss of baby (deciduous) teeth, but no bone abnormalities. TNALP is expressed in the liver, kidney and bone, and its substrates include TNSALP inorganic pyrophosphate, pyridoxal-5′-phosphate (PLP) / vitamin B6, and phosphoethanolamine (PEA). Major hallmarks of HPP are low alkaline phosphatase (ALP) and elevated PLP PEA, and inorganic pyrophosphate (PPi) levels. PPi is an inhibitor of mineralization that controls mineral entry into the skeleton. Elevated PPi levels can block calcium and phosphorus from entering bone, and thereby cause elevated levels of calcium in the blood and urine. Generally, HPP severity correlates with how much alkaline phosphatase activity remains in the body, with less enzyme activity causing more severe disease. In 2015, the U.S. Food and Drug Administration (FDA) approved the enzyme replacement therapy asfotase alfa (Strensiq), a TNALP produced by recombinant DNA technology in a Chinese hamster ovary cell line, as the first medical treatment for perinatal, infantile and juvenile-onset HPP. Asfotase alfa is a soluble glycoprotein composed of two identical polypeptide chains. Each chain contains 726 amino acids with a theoretical mass of 161 kDa. Each chain consists of the catalytic domain of human TNALP, the human immunoglobulin G1 Fc domain and a deca- aspartate peptide used as a bone targeting domain. The two polypeptide chains are covalently linked by two disulfide bonds. In the United States, patients of any age with pediatric-onset HPP are eligible for this bone-targeted TNALP replacement therapy given by subcutaneous injection either 3 times per week (2 mg / kg) or 6 times per week (1 mg / kg). While the use of asfotase alfa is effective for treatment, especially for infantile HPP, many questions regarding its long-term effects, the management of treatment, and any potential secondary adverse effects remain unresolved. For example, Strensiq contains a black-box warning relating to hypersensitivity reactions to asfotase alfa, including anaphylaxis. In addition, the use of asfotase alfa has been associated with ectopic calcification attributed to reductions in circulating PPi, allowing the propagation of hydroxyapatite crystals—including outside of skeletal sites such as in the eye and in the kidneys. Most importantly, however, patients administered asfotase alfa may develop anti-asfotase alfa antibodies, leading to immune-mediated loss of effectiveness (LoE) and the potential for disease progression. For example, among Strensiq-treated patients with HPP in clinical studies who had post-baseline antibody data available, 97 / 109 (89%) tested positive for anti-asfotase alfa antibodies at some time point during treatment. Among those 97 patients, 55 (57%) also tested positive for neutralizing antibodies at some time point during Strensiq treatment. The use of immunosuppressive therapy (IST) in patients treated with asfotase alfa who demonstrate LoE is currently being investigated. Gene therapy is an innovative approach in medicine aimed at treating inherited and acquired diseases through the delivery of new genetic material into cells of a patient to compensate for or suppress the function of a mutant gene and / or treat a disorder, and has been suggested as a potential approach for treating HPP. Recent advances have seen a number of gene therapies approved for marketing in the United States. The approved therapies treat a range of disorders, including but not limited to sickle cell disease, Duchenne muscular dystrophy, inherited retinal disorder, hemophilia B, and others. The majority of vectors used in approved gene therapies are viral, in which AAV is the most widely used viral vector for in vivo gene therapy applications (Zhao et al., Viral vector‐based gene therapies in the clinic, Bioeng Transl Med. 2022 Jan; 7(1): e10258). The primary reasons for using viral vectors for gene delivery include their natural ability to infect target cells, efficient shuttling of genetic materials into the nucleus of target cells, and, where desirable, insertion into the target cell genome (see, e.g., Finer et al., A brief account of viral vectors and their promise for gene therapy. Gene Ther.2017;24(1):1‐2). Use of viral vectors, such as AAV for example, have limited use in clinical practice due to the immune response generated to the viral vector, which presents a significant hurdle to the translation of viral vector‐based gene therapies. The inherent problems associated with the use of viral vectors in gene therapies have led researchers to examine alternatives, including non-viral vectors. Because it is far more difficult for non-viral vectors (RNA or DNA) to transfect many specific cell types compared to viral vectors, the DNA or RNA expression cassette is often complexed with delivery vehicles (e.g., cationic lipids, cationic polymers, etc.) or subjected to forced entry (e.g., electroporation, hydrodynamic injection, etc.). Potential advantages of using non-viral vectors for gene therapies include larger cargo packing capacity compared to, for example, AAV and LVV, thus expanding the potential disorders that can be targeted due to the ability to encode for larger genes. Furthermore, non-viral vectors are generally less immunogenic than viral vectors (see Hardee et al., Advances in Non- Viral DNA Vectors for Gene Therapy. Genes 2017, 8(2), 65). Non-viral vectors, however, have historically been beset with some notable inherent limitations resulting in limited clinical success to date. Because plasmids are non-replicating episomes, transgene expression is transient and diluted by cell division. Non-viral vectors also tend to achieve lower rates of gene delivery compared to select viral vectors. This lower efficiency, especially with DNA-based vector systems, is largely due to the difficulty of getting the expression cassette into the nucleus of target cells, which is classically considered as one of the main bottlenecks of non-viral plasmid-based expression systems (Puras et al., Protamine / DNA / Niosome Ternary Nonviral Vectors for Gene Delivery to the Retina: The Role of Protamine. Mol. Pharm. 2015, 12, 3658–3671). Furthermore, early generation plasmids typically included antibiotic resistance-encoding genes for selection of plasmid-harboring bacteria during production. The use of antibiotics and their resistance genes in the preparation of plasmid vectors, however, is discouraged by regulatory bodies such as the Food and Drug Administration and the European Medicines Agency because of the risk of transfer and replication of resistance genes to bacteria in the human microbiome and possibly into the environment. Additionally, residual antibiotics that remain from vector production may trigger an immune reaction in patients. Minimized plasmid DNA have recently been developed to mitigate some disadvantages associated with the use of conventional plasmids. For example, Aldevron LLC and Nature Technology Corporation have developed minicircle DNA plasmids known as Nanoplasmids®. These plasmids were ultimately developed after removing all non-essential sequences from the plasmid backbone, including extraneous bacterial DNA flanking the selection marker and replication origin (Ori), bacterial sequences resulting in the formation of secondary structures (e.g., Z DNA, cruciforms, palindromes, repeats), cryptic splice sites, sequences resulting in RNA secondary structures, sequences with human genome homology, alternative reading frames, cryptic promoters and chi sites (see Hodgson et al., Recent advances in non-viral vectors for gene therapy & vaccination. Cell Gene Therapy Insights 2017; 3(2), 95-101). In addition, the antibiotic resistance genes in the bacterial backbone were replaced with a small RNA selectable marker known as an RNA-Out, which allows selection with sucrose rather than antibiotics (see id.). The plasmids were further optimized to use an R6K-derived bacterial Ori, a smaller bacterial origin of replication than those traditionally used in plasmids such as the pUC Ori(id.). These modifications result in a plasmid with a bacterial Ori and selection backbone of less than 0.5 kb (id.). Such small, circular plasmids have also recently been implemented in the production of AAV viral-vectors and have shown improved production capacities (see, e.g., WO2019 / 183248). As noted above, one particular problem in using non-viral DNA vectors in gene therapies has been the inability to achieve long-term persistence of gene expression, such as that attained with AAV viral vectors or integrating lentivirus vectors. Recently, non-viral delivery of linear DNAs having modified geometry such as closed-end linear duplex DNA (CELiD) have shown promise as an alternative to viral vectors due to increased transgene expression and less cytotoxicity. CELiDs consists of double-stranded DNA molecules with covalently closed terminal hairpins (see, e.g., Li et al., Production and characterization of novel recombinant adeno-associated virus replicative-form genomes: A eukaryotic source of DNA for gene transfer. PLoS ONE 8, e69879 (2013)). A particular-type of CELiD is generated as an intermediate of AAV replication in eukaryotic cells, and such structures are in effect a double stranded AAV genome containing a DD- ITR (“double-D”) element capped with hairpin-forming palindromic terminal regions (see, e.g., Samulski et al., AAV-mediated gene therapy for research and therapeutic purposes. Annu. Rev. Virol.1, 427–451 (2014); see also US2021 / 0269828). However, the question of whether modified- end linear DNAs can also provide a safe means of non-integrating gene transfer remains unanswered, as such linear DNA structures have concerningly shown a tendency to stably transfect high fractions of cells in vivo, sometimes as high as between 10 and 20% of the initially transfected cells (Lim et al., High spontaneous integration rates of end-modified linear DNAs upon mammalian cell transfection. Sci Rep 13, 6835 (2023)). Given the issues associated with the use of asfotase alfa (Strensiq), improved HPP treatments are needed to adequately meet the therapeutic requirements of those suffering from this disorder. SUMMARY OF THE DISCLOSURE The present disclosure provides compositions, and methods of using such composition, comprising isolated, circular, non-integrating, non-viral DNA vectors having improved expression of TNALP, wherein the DNA vectors are capable of forming one or more specialized secondary structures, for example one or more extended cruciform structures, to treat a human with HPP. The DNA vectors as described herein comprise novel TNALP expression constructs and a specialized secondary structure, for example at least one or more extended cruciform structures, wherein the novel TNALP expression construct and specialized secondary structure in combination provide exceptional stability and reduced immunogenicity, leading to elevated TNALP transgene activity that is durably maintained for an extended length of time (see, e.g., FIG. 2), leading to increased potency in a subject (see, e.g., Example 2). The DNA vectors as described herein are capable of being repeatedly re-dosed, thus providing significant improvements over viral vector approaches. In one aspect, provided herein is a DNA vector comprising: (i) a first portion comprising: a) a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 1, wherein the nucleic acid is selected from SEQ ID NOS: 2, 3, or 4, or a nucleic acid at least about 95% identical thereto; b) a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 5, wherein the nucleic acid is SEQ ID NO: 6, or a nucleic acid at least about 95% identical thereto; or, c) a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 7, wherein the nucleic acid is SEQ ID NO: 8, or a nucleic acid at least about 95% identical thereto; d) a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 9, wherein the nucleic acid is SEQ ID NO: 10, or a nucleic acid at least about 95% identical thereto; or, e) a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 11, wherein the nucleic acid is SEQ ID NO: 12, or a nucleic acid at least about 95% identical thereto (ii) a second portion comprising a nucleic acid sequence capable of forming at least one specialized secondary structure, for example an extended cruciform structure, wherein the second portion includes two inverted terminal repeat (ITR) sequences and a non-repeating sequence contained between the two repeating sequences, wherein the ITRs are capable of aligning to form two double-stranded arms of significant length (e.g., at least about 80 bps to about 110 bps or longer), with a loop formed at the end of each extended arm by the non-repeating sequence, wherein the non-repeating sequence is between about 225 bps and about 460 bps and comprises a bacterial origin of replication or fragment thereof, and wherein the second portion lacks a bacterial selection marker. In some embodiments, the second portion of the DNA vector comprises a nucleic acid sequence having any one of SEQ ID NOS: 13, 14, 15, or 16, or a sequence at least 95% identical thereto. In some embodiments, the DNA vector has a nucleic acid sequence selected from SEQ ID NOS: 17, 18, 19, 20, 21, 22, or 23, or a DNA vector having a nucleic acid sequence at least about 95% identical thereto. In alternate embodiments, the DNA vector has a nucleic acid sequence selected from SEQ ID NOS: 97 or 98, or a DNA vector having a nucleic acid sequence at least about 95% identical thereto. In some embodiments, the first portion of the DNA vectors provided herein express a TNALP polypeptide comprising the formula [A]–[B]–[R]–[E]y, wherein: [A] comprises a secretion signal peptide comprising the amino acid sequence of SEQ ID NO: 24, wherein the amino acid is encoded by a nucleic acid sequence of SEQ ID NO: 25, or a nucleic acid sequence at least about 95% identical thereto; [B] comprises human TNALP, wherein the TNALP is selected from the amino acid sequence of SEQ ID NO: 26, wherein the amino acid is encoded by a nucleic acid sequence of SEQ ID NO: 27, or a nucleic acid sequence at least about 95% identical thereto, or a human TNALP derivative having an E108M substitution, the TNALP derivative having the amino acid sequence of SEQ ID NO: 28, wherein the amino acid is encoded by a nucleic acid sequence of SEQ ID NO: 29, or a nucleic acid sequence at least about 95% identical thereto; [R] is –(M(Fc)N)–, wherein M is a dipeptide selected from -L-K-, -L-S-, or -S-S-, wherein N is the dipeptide D-I, wherein Fc is an Fc domain having an amino acid sequence of SEQ ID NO: 30, and is encoded by the nucleic acid sequence of SEQ ID NO: 31, or a nucleic acid sequence at least about 95% identical thereto; and, [E] comprises the bone targeting amino acid sequence -D-S-S-, wherein y = 6 (SEQ ID NO: 32). In some embodiments, the first portion of the DNA vectors provided herein express a TNALP polypeptide comprising the formula [A]–[B]–[R]–[E]y, wherein: [A] comprises a secretion signal peptide comprising the amino acid sequence of SEQ ID NOS: 24, 74, 75, 76, 77, 78, 79, 80, 81, 82, or 83; [B] comprises human TNALP, wherein the TNALP is selected from the amino acid sequence of SEQ ID NO: 26, wherein the amino acid is encoded by a nucleic acid sequence of SEQ ID NO: 27, or a nucleic acid sequence at least about 95% identical thereto, or a human TNALP derivative having an E108M substitution, the TNALP derivative having the amino acid sequence of SEQ ID NO: 28, wherein the amino acid is encoded by a nucleic acid sequence of SEQ ID NO: 29, or a nucleic acid sequence at least about 95% identical thereto, or a human TNALP derivative having the amino acid sequence of SEQ ID NO: 85, 86, 87, 88, 89, 90, 91, 95, or 96; [R] is –(M(Fc)N)–, wherein M is a dipeptide selected from -L-K-, -L-S-, or -S-S-, wherein N is the dipeptide D-I, wherein Fc is an Fc domain having an amino acid sequence of SEQ ID NO: 30, and is encoded by the nucleic acid sequence of SEQ ID NO: 31, or a nucleic acid sequence at least about 95% identical thereto; and, [E] comprises the bone targeting amino acid sequence -D-S-S-, wherein y = 6 (SEQ ID NO: 32). In some embodiments, the nucleic acids of the first portion encoding the TNALP polypeptide comprise improved promoter and / or enhancer element DNA sequences, for example a zero-CpG promoter and / or zero-CpG enhancer element driving expression of the TNALP polypeptide. The improved organization of the promoter and / or enhancer elements of the DNA vectors described herein result in robust expression of TNALP such that ALP is measured at consistently high levels in plasma in vivo over several weeks (see, e.g., FIG. 2). It is further believed that the use of a zero-CpG promoter and / or zero-CpG enhancer assists in further reducing epigenetic downregulation of transgene expression in vivo by reducing methylation of the promoter / enhancer region of the DNA construct. In addition, the use of particular enhancer elements structured as provided herein may further assist in the recruitment of RNA polymerase, increasing transcription of the TNALP transgene of the DNA construct. In some embodiments, the DNA construct includes a zero-CpG promoter, that is, the promoter does not include, in a 5’ to 3’ direction, CpG dinucleotides. In some embodiments, the DNA construct includes a zero-CpG enhancer element. In some embodiments, the DNA construct includes both a zero-CpG promoter and a zero-CpG enhancer element. In some embodiment, the zero-CpG promoter is a EF1 alpha long (EF1L) promoter, e.g., a promoter having the nucleic acid sequence of SEQ ID NO: 33, or a nucleic acid sequence at least 95% identical thereto. In some embodiments, an EFIL promoter having the nucleic acid sequence of SEQ ID NO: 34, or a nucleic acid sequence at least about 95% identical thereto, drives the expression of the TNALP polypeptide. In some embodiments, the EF1L promoter is amplified by an ApoE enhancer element, e.g., an enhancer element having the nucleic acid sequence of SEQ ID NO: 35, or a nucleic acid sequence at least 95% identical thereto. The presence of a constant region of the human IgG1 Fc domain in the TNALP polypeptide increases long term ALP plasma levels, thus enhancing the potency of the DNA vector. The TNALP polypeptide includes an Fc domain linked to a first hinge (i.e., “M” as described herein) on the N-terminus of the Fc domain and a second hinge (i.e., “N” as described herein) on the C- terminus of the Fc domain. The bone targeting peptide (DSS)6 is further fused to the C-terminus of the second hinge. The fusing of an Fc domain to the TNALP increases activity of TNALP, as the first hinge increases flexibility of the polypeptide encoded by the expression cassette. In some embodiments, the first hinge or “M” of the DNA vectors described herein comprises the dipeptide -L-K-, -L-S-, or -S-S-. In some embodiments, the first hinge comprises the dinucleotide -L-K-. In some embodiments, the first hinge comprises the dinucleotide -L-S-. In some embodiments, the first hinge comprises the dinucleotide -S-S-. The second hinge offers further optimization of the encoded polypeptide such that the Fc domain is flexibly conjugated to the bone-targeting peptide (for example, the (DSS)6 peptide (SEQ ID NO: 32) to guide TNALP to the area in need in the subject to be treated, for example a human having hypophosphatasia (HPP), a bone mineralization disorder. In some embodiments, the second hinge or “N” of the DNA vectors described herein comprises the dipeptide -D-I-. Furthermore, the robust potency and stability of the DNA vectors described herein can also be attributed to a reduced immunogenicity of the vector due to, in addition to the inclusion of a specialized secondary structure as described further below, several nucleic acid components of the improved expression cassette being further designed to be devoid of CpG motifs (i.e., zero-CpG or “zg”). For example, the DNA vectors described herein comprise one or more of the following elements that are zero-CpG, including: a zg-EF1L promoter nucleic acid sequence, a zg-TNALP nucleic acid sequence, a zg-Fc domain nucleic acid sequence, and a zg-bone targeting peptide encoding nucleic acid sequence. In addition, further expression elements may be operatively arranged 3’ downstream of the bone-targeting peptide encoding sequence. For example, in some embodiments, the DNA vector comprises a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE) (e.g., WPRE2) and / or bGH2 poly(A) polyadenylation sequence. These elements further contribute to efficient transcription and expression of the transgene which lead to outstanding TNALP expression in a subject. The second portion of the DNA vectors provided herein comprise a nucleic acid sequence that is capable of forming a specialized secondary structure, for example, at least one or more extended cruciform structures. In alternative embodiments, the specialized structure may take the form of another type of non-canonical B-DNA forming secondary structure like a hairpin, G- quadruplex, or i-motif. The specialized secondary structure, for example an extended cruciform structure, contributes to increased nucleus entry, episomal stability, and long-term transgene expression, and provides reduced immunogenicity and epigenetic downregulation. The specialized secondary structure is formed through the interaction of two specifically oriented repeating sequences, for example AAV-derived inverted terminal repeat (ITR) sequences, separated by a small, non-repeating sequence, for example a non-repeating sequence comprising a small bacterial origin of replication (ori). By specifically orienting these two repeating sequences to flank a non-repeating sequence of particular size, for example an ori of between about 225 and 460 base-pairs (bps), the two repeating sequences are capable of aligning to form, for example, an extended cruciform structure containing a Holliday junction and two double-stranded arms of significant length (e.g., 80-110 bps or more), with a loop formed at the end of each extended arm by the non-repeating sequence. Importantly, the loop capable of being formed at the end of each extended arm by the non-repeating sequence may also form additional secondary structures such as small hairpins, adding to the stability of the DNA vector. Comparatively, using a single ITR in a circular DNA vector, for example a DD-ITR, or failing to sufficiently spatially orient the repeating sequences using an appropriately sized non-repeating sequence, results in one or more hairpin-like structures or short cruciform structures with shorter aligned arms (e.g., less than 50 bps). Accordingly, significant advantages in nucleus entry, transgene expression and persistence, and reduced immunogenicity are achieved with the circular DNA vectors of the present invention containing unique transgene expression cassettes and specialized secondary structures, for example an extended cruciform structure, making these DNA vectors particularly useful in treating humans with HPP. While not wanting to be limited to one theory, it is proposed that the capability to form specialized secondary structure, for example an extended cruciform structure, mimics endogenous secondary DNA structures (e.g., cruciform and Holliday junctions), which can bind to nuclear import proteins to facilitate the transcription of a transgene while not provoking a toxic inflammatory response or inducing epigenetic downregulation. The lack of the development of an adaptive immune response to the DNA-vectors described herein is important for three critical reasons. First, it allows for repeat dosing of the DNA vectors described herein as needed. Second, it indicates that the DNA vectors described herein may not be subjected to gene silencing in vivo, a critically limiting aspect of prior art viral and non-viral vectors. Finally, by retaining a bacterial origin of replication without inducing unwanted immunogenicity, the ability of the DNA vectors to be manufactured in a suitable bacterial host is preserved, significantly reducing production costs. Importantly, the extended cruciform DNA vectors provided herein do not comprise or form an AAV ITR-DD element, that is an ITR having both a D element and D’ element in the inverted terminal repeat (a “double-D ITR”), as commonly observed with recombinant AAV vectors in vivo (see, e.g., Schnepp et al., Recombinant Adeno-Associated Virus Vector Genomes Take the Form of Long-Lived, Transcriptionally Competent Episomes in Human Muscle. Hum Gene Ther.2016 Jan 1; 27(1): 32–42). Such vectors having a double-D ITR may have increased immunogenicity and decreased transgene expression and persistence compared to the DNA vectors described herein. As described above, the ITR sequences are separated by a non-repeating sequence of appropriate length, which may allow for the alignment of the ITR sequences and the formation of double-stranded arms of sufficient length—for example at least about 80 bps, at least about 90 bps, at least about 100 bps, or at least about 110 bps—in the specialized secondary structure. In some embodiments, the ITR sequences are derived from one or more AAV serotypes, for example, but not limited to, AAV1, AAV2, AAV3, AAV4, AAV5, AVV6, AVV7, AAV8, or AAV9. Suitable AAV ITR sequences for use in the DNA vectors described herein include any of SEQ ID NOS: 36-53, or a sequence at least about 85%, at least about 90%, or at least about 95% identical thereto. As provided herein, the repeating sequences of the extended cruciform-forming DNA vectors described herein flank a non-repeating sequence of between about 225 bps and about 460 bps, wherein the non-repeating sequence comprises a bacterial ori and lacks a bacterial selection marker. The non-repeating sequence provides appropriate spacing for the ITR sequences, allowing the ITR sequences to appropriately align to form, for example, double-stranded extended arms of a cruciform, with the non-repeating sequence forming a loop between the aligned repeating sequences at the top of each extended arm. In some embodiments, the DNA sequence elements that lead to the formation of, for example, the extended cruciform DNA structure may contain additional secondary structure that stabilizes the formation of the larger cruciform structure. Thus, in some embodiments, the non-repeating sequence may contribute additional structure to the DNA vector, thus further enhancing the stability of the DNA vector. In some embodiments, the non- repeating sequence comprises a bacterial origin of replication (ori) having a sequence of between about 225 bps and about 400 bps. In some embodiments, the non-repeating sequence comprises an ori having a sequence of less than about 400 bps. In some embodiments, the non-repeating sequence comprises an ori having a sequence of less than about 350 bps. In some embodiments, the non-repeating sequence comprises an ori having a sequence of less than about 300 bps. In some embodiments, the non-repeating sequence comprises an ori of between about 375 bps and about 400 bps. In some embodiments, the non-repeating sequence comprises an ori of between about 290 bps and about 310 bps. In some embodiments, the non-repeating sequence comprises an ori of between about 235 bps and about 260 bps. In some embodiments, the ori is derived from pR6K (see, e.g., Rakowski et al., Plasmid R6K Replication Control. Plasmid.2013 May; 69(3): 231–242, incorporated herein by reference). In some embodiments, the ori is derived from R6Kγ. In some embodiments, the non-repeating sequence comprises an ori selected from SEQ ID NOS: 54, 55, 56, 57, or 58, or a sequence at least about 85%, at least about 90%, or at least about 95% identical thereto. In some alternative embodiments, the ori is derived from pMB1, pBR322, ColE1, p15A, pSC101, or F1. In some embodiments, in addition to an ori sequence, the non-repeating sequence may contain additional DNA sequences, for example, small runs of extraneous and or spacer nucleotide sequences (e.g., from about 1 bp to about 20 bps), cloning or recombination sites, and / or other sites such as LoxP sites, FRT sites, attB and attP sites or their product sites attL or attR, or alternative recombination target sites derived from these sites, e.g., Lox511 or Lox66 sites. The non-repeating sequence comprising an ori does not include a bacterial selection marker, for example, a drug resistance gene or an RNA-based selectable marker such as an RNA-IN or RNA- Out selectable marker. In some embodiments, the second portion of the DNA vector comprises a first ITR nucleic acid sequence having any one of SEQ ID NOS: 36, 38, 40, 42, 44, 46, 48, 50, or 52, or a sequence at least about 85%, at least about 90%, or at least about 95% identical thereto; a non-repeating nucleic acid sequence comprising an ori having any one of SEQ ID NOS: 54, 55, 56, 57, or 58, or a sequence at least about 85%, at least about 90%, or at least about 95% identical thereto; and a second ITR nucleic acid repeating sequence having any one of SEQ ID NOS: 37, 39, 41, 43, 45, 47, 49, 51, or 53, or a sequence at least 95% identical thereto. In some embodiments, the second portion of the DNA vector comprises a nucleic acid sequence having any one of SEQ ID NOS: 13, 14, 15, or 16, or a sequence at least about 85%, at least about 90%, or at least about 95% identical thereto. In some embodiments, the extended cruciform DNA vector has the nucleic acid sequence of SEQ ID NO: 17, or a nucleic acid sequence having at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 17 is a human TNALP (hTNALP)-encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an ApoE-EF1L-driven zg-hTNALP-zg-Fc-(DSS)6 transgene further comprising a woodchuck hepatitis virus post-transcriptional regulatory element 2 (WPRE2) and bGH2 poly(A) polyadenylation sequence. In some embodiments, the extended cruciform DNA vector has the nucleic acid sequence of SEQ ID NO: 18, or a nucleic acid sequence having at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 18 is an hTNALP polypeptide-encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) a zg-EF1L-zg-driven hTNALP-zg-Fc-(DSS)6 transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The TNALP polypeptide includes a secretion signal. In some embodiments, the extended cruciform DNA vector has the nucleic acid sequence of SEQ ID NO: 19, or a nucleic acid sequence having at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 19 is an hTNALP-encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an ApoE-zg- EF1L-zg-driven hTNALP-zg-Fc-(DSS)6 transgene further comprising WPRE2 and a bGH2 poly(A) polyadenylation sequence. The TNALP polypeptide includes a secretion signal. In some embodiments, the extended DNA vector has the nucleic acid sequence of SEQ ID NO: 20, or a nucleic acid sequence having at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 20 is an hTNALP-encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an EF1L-zg- hTNALP-hinge2-zg-Fc-(DSS)6transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The TNALP polypeptide includes a secretion signal. In some embodiments, the extended DNA vector has the nucleic acid sequence of SEQ ID NO: 21, or a nucleic acid sequence having at least 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 21 is an hTNALP-encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an EF1L-driven zg- hTNALP-hinge3-zg-Fc-(DSS)6 transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The TNALP polypeptide includes a secretion signal. In some embodiments, the extended DNA vector has the nucleic acid sequence of SEQ ID NO: 22, or a nucleic acid sequence having at least 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 22 is an hTNALP-derivative encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an zg-EF1L- driven zg-hTNALP(E108M)-hinge1-zg-Fc-(DSS)6 transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The TNALP derivative comprises an E108M amino acid substitution in the active site of the TNALP and a secretion signal. In some embodiments, the extended DNA vector has the nucleic acid sequence of SEQ ID NO: 23, or a nucleic acid sequence having at least 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 23 is an hTNALP-derivative encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an EF1L-driven zg-hTNALP(E108M)-hinge3-zg-Fc-(DSS)6 transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The TNALP derivative comprises an E108M amino acid substitution in the active site of the TNALP and a secretion signal. In alternate embodiments, the extended DNA vector has the nucleic acid sequence of SEQ ID NO: 97, or a nucleic acid sequence having at least 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 97 is an hTNALP-derivative encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an EF1L-driven zg-hTNALP(N213Q, N286Q)-hinge3-zg-Fc-(DSS)6 transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The TNALP derivative comprises a N213Q and a N286Q amino acid substitution in the active site of the TNALP and a secretion signal. In alternate embodiments, the extended DNA vector has the nucleic acid sequence of SEQ ID NO: 98, or a nucleic acid sequence having at least 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 98 is an hTNALP-derivative encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an EF1L-driven zg-hTNALP(E108M, N213Q, N286Q)-hinge3-zg-Fc-(DSS)6 transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The TNALP derivative comprises an E108M , a N213Q, and a N286Q amino acid substitution in the active site of the TNALP and a secretion signal. In an alternative aspect, the first portion of the DNA vectors described herein alternatively comprise a nucleic acid sequence encoding a TNALP polypeptide comprising the formula [A]– [B]–[R]–[E]y, wherein: [A] comprises a secretion signal peptide; [B] comprises human TNALP, wherein the TNALP is selected from a TNALP having the amino acid sequence of SEQ ID NO: 26, wherein the amino acid is encoded by a nucleic acid sequence of SEQ ID NO: 27, or a nucleic acid sequence at least about 85%, at least about 90%, or at least about 95% identical thereto, or a human TNALP derivative having an E108M substitution, the TNALP derivative having the amino acid sequence of SEQ ID NO: 28, wherein the amino acid is encoded by a nucleic acid sequence of SEQ ID NO: 29, or a nucleic acid sequence at least about 85%, at least about 90%, or at least about 95% identical thereto, or a human TNALP having the amino acid sequence of SEQ ID NO: 26 further comprising one or more of the following substitutions: E108X, N213X, or N286X, wherein X = any amino acid (SEQ ID NO: 91), or a human TNALP derivative having the amino acid sequence of SEQ ID NO: 85, 86, 87, 88, 89, 90, 95, or 96; [R] is –(M(Fc)N)–, wherein M is the dipeptide -L-S- or -S-S-, wherein N is the dipeptide D-I, wherein Fc is an Fc domain; and, [E] comprises a bone targeting amino acid sequence, wherein y = 3-30; wherein the first portion further comprises a nucleic acid sequence comprising a promoter operably linked to the nucleic acid encoding the TNALP polypeptide; and, (i) a second portion comprising a nucleic acid sequence capable of forming at least one specialized secondary structure, for example an extended cruciform structure, wherein the second portion includes two inverted terminal repeat (ITR) sequences and a non-repeating sequence contained between the two repeating sequences, wherein the ITRs are capable of aligning to form two double-stranded arms of significant length (e.g., at least about 80 bps - about 110 bps or longer), with a loop formed at the end of each extended arm by the non-repeating sequence, wherein the non-repeating sequence is between about 225 bps and about 460 bps and comprises a bacterial origin of replication or fragment thereof, and wherein the second portion lacks a bacterial selection marker. In some embodiments, M is the dipeptide -L-S-. In some embodiments, M is the dipeptide -S-S-. It has been found that the use of novel hinge domains, for example -L-S-Fc-D-I- (SEQ ID NO: 92) and -S-S-Fc-D-I- (SEQ ID NO: 93), separating the hTNALP peptide and the Fc domain- encoding sequence further increase flexibility and efficacy of the encoded polypeptide. The bone targeting amino acid sequence can be any suitable bone targeting amino acid sequence, including, for example, aspartate (D), glutamate (E), aspartate-serine-serine (DSS), glutamate-glutamate- serine (EES), and valine-histidine-histidine (VHH). In some embodiments, the bone targeting amino acid sequence [E] is aspartate. In some embodiments, the bone targeting amino acid sequence [E] is glutamate. In some embodiments, the bone targeting amino acid sequence [E] is poly(aspartate-serine-serine (DSS)). In some embodiments, the bone targeting amino acid sequence [E] is glutamate-glutamate-serine (EES). In some embodiments, the bone targeting amino acid sequence [E] is aspartate-serine-serine (DSS). In some embodiments, [E] is (DSS) and y = 6 (SEQ ID NO: 32). In some embodiments, the Fc domain comprises an amino acid sequence of SEQ ID NO: 30, and is encoded by the nucleic acid sequence of SEQ ID NO: 31, or a nucleic acid at least about 85%, at least about 90%, or at least about 95% identical thereto. In an alternative aspect, the first portion of the DNA vectors described herein alternatively comprise a nucleic acid sequence encoding a TNALP polypeptide comprising the formula [A]– [B]–[R]–[E]y, wherein: [A] comprises a secretion signal peptide; [B] comprises human TNALP, wherein the TNALP is selected from a human TNALP derivative having an E108M substitution, the TNALP derivative having the amino acid sequence of SEQ ID NO: 28, wherein the amino acid is encoded by a nucleic acid sequence of SEQ ID NO: 29, or a nucleic acid sequence at least about 85%, at least about 90%, or at least about 95% identical thereto, or a human TNALP derivative having the amino acid sequence of SEQ ID NO: 26, and further comprising one or more of the following substitutions: E108X, N213X, or N286X, wherein X = any amino acid (SEQ ID NO: 91), or a human TNALP derivative having the amino acid sequence of SEQ ID NO: 85, 86, 87, 88, 89, 90, 95, or 96; [R] is –(M(Fc)N)–, wherein M is the dipeptide -L-K-, -L-S- or -S-S-, wherein N is the dipeptide D-I, wherein Fc is an Fc domain; and, [E] comprises a bone targeting amino acid sequence, wherein y = 3-30; wherein the first portion further comprises a nucleic acid sequence comprising a promoter operably linked to the nucleic acid encoding the TNALP polypeptide; and, (ii) a second portion comprising a nucleic acid sequence capable of forming at least one specialized secondary structure, for example an extended cruciform structure, wherein the second portion includes two inverted terminal repeat (ITR) sequences and a non-repeating sequence contained between the two repeating sequences, wherein the ITRs are capable of aligning to form two double-stranded arms of significant length (e.g., at least about 80 bps - about 110 bps or longer), with a loop formed at the end of each extended arm by the non-repeating sequence, wherein the non-repeating sequence is between about 225 bps and about 460 bps and comprises a bacterial origin of replication or fragment thereof, and wherein the second portion lacks a bacterial selection marker. In some embodiments, M is the dipeptide -L-K-. In some embodiments, M is the dipeptide -L-S-. In some embodiments, M is the dipeptide -S-S-. In some embodiments, the human TNALP derivative having the amino acid sequence of SEQ ID NO: 26, further comprises an E108M substitution (SEQ ID NO: 28). In some embodiments, the human TNALP derivative having the amino acid sequence of SEQ ID NO: 26, further comprises a N213Q substitution (SEQ ID NO: 95). In some embodiments, the human TNALP derivative having the amino acid sequence of SEQ ID NO: 26, further comprises a N286Q mutation (SEQ ID NO: 96). In some embodiments, the human TNALP derivative having the amino acid sequence of SEQ ID NO: 26, further comprises a E108M substitution, N213Q substitution, and N286Q substitution (SEQ ID NO: 88). The bone targeting amino acid sequence can be any suitable bone targeting amino acid sequence, including, for example, aspartate (D), glutamate (E), aspartate-serine-serine (DSS), glutamate-glutamate- serine (EES), and valine-histidine-histidine (VHH). In some embodiments, the bone targeting amino acid sequence [E] is aspartate. In some embodiments, the bone targeting amino acid sequence [E] is glutamate. In some embodiments, the bone targeting amino acid sequence [E] is aspartate-serine-serine (DSS). In some embodiments, the bone targeting amino acid sequence [E] is poly (glutamate-glutamate-serine (EES)). In some embodiments, the bone targeting amino acid sequence [E] is aspartate-serine-serine (DSS). In some embodiments, [E] is (DSS) and y = 6 (SEQ ID NO: 32). In some embodiments, the Fc domain comprises an amino acid sequence of SEQ ID NO: 30, and is encoded by the nucleic acid sequence of SEQ ID NO: 31, or a nucleic acid at least about 85%, at least about 90%, or at least about 95% identical thereto. Also provided herein are pharmaceutical compositions comprising a DNA vector having an improved TNALP expression cassette as described herein in a pharmaceutically acceptable medium, for example, wherein the DNA vector is formulated with one or more lipid nanoparticles (LNP). In some embodiments, the DNA vector is formulated with one or more polymer nanoparticles. In some embodiments, the DNA vector is formulated with one or more proteo-lipid nanoparticles. In some embodiments, the DNA vector is formulated with one or more microbubbles. In some embodiments, the DNA vector is administered as a pharmaceutical composition formulated with a natural or synthetic membrane or an implanted vehicle which improves the therapeutic activity of the DNA vector by more efficient delivery. In some embodiments, the DNA vector is formulated in an implanted delivery vehicle. In some embodiments, the implanted delivery vehicle comprises a polymeric hydrogel. In some embodiments, the implanted delivery vehicle comprises a porous or permeable membrane. In some embodiments, the DNA vector may be given to the patient with physical methods, such as electroporation, sonoporation with microbubbles, sonoporation without microbubbles, magnetofection, hydroporation, photoporation, mechanical massage, jet injection, biolistics (gene gun), hydrodynamic injection, needle injection, or microinjections. BRIEF DESCRIPTION OF THE FIGURES For a general understanding of the features of the disclosure, reference is made to the drawings. In the drawings, like reference numerals have been used throughout to identify identical elements. FIG. 1A illustrates an exemplary circular, non-integrating, non-viral DNA vector having an improved TNALP expression cassette as described herein comprising (i) a first portion comprising an expression cassette encoding a TNALP polypeptide, where the nucleic acid sequence encoding the TNALP polypeptide is operatively linked at least to a promoter, and the TNALP polypeptide comprises, from the N-terminus to C-terminus, a secretory signal, TNALP, a first hinge region, an Fc domain, a second hinge region, and a bone targeting peptide tag; and (ii) a second portion capable of forming one or more specialized secondary structures, for example an extended cruciform structure, wherein the second portion comprises at least about two inverted repeat sequences, wherein the at least about two inverted repeat sequences are separated by a non- repeating nucleotide sequence comprising a bacterial Ori of between about 225 and about 460 base-pairs. FIGS. 1B and 1C illustrate the DNA vector of FIG. 1A and illustrate a specialized secondary structure in the form of, for example, an extended cruciform structure as described herein. The DNA vector’s second portion contains a nucleic acid sequence that may be capable of forming the extended cruciform structure comprising repeat elements that lead to the formation of the extended cruciform DNA structure and a non-repeating sequence. The repeat elements and non-repeating elements may contain additional secondary structure that stabilizes the formation of the larger cruciform structure. As depicted in FIG.1B, the cruciform structure is formed between specific repeat elements, however, due to the nature of these repeats, the cruciform may be formed between other repeat elements, or between the inverted repeat sequences and the remainder of the non-viral DNA vector. In some embodiments, the inverted repeat elements may also be discontinuous, with regions of non-base paired or single-stranded DNA. One such region containing non-base paired or single-stranded DNA is in the region comprising the non-repeating nucleotide sequence, for example a bacterial Ori sequence, having at least about between about 25 and about 460 base-pairs, which is internal to the cruciform structure (see FIG. 1A). Given the components of the DNA vectors of the present disclosure, it is believed that at least about two inverted repeat sequences, each separated by a non-repeating nucleotide sequence having at least about between about 25 and 460 base-pairs, facilitates the formation of a specialized structure, for example a single extended cruciform structure having two long arms capped by a loop region of single-stranded DNA. FIG. 2 illustrates the persistence of the DNA vectors described herein encoding human TNALP transgenes (M080 – SEQ ID NO: 17; M086 – SEQ ID NO: 18; M088 – SEQ ID NO: 19; M089 – SEQ ID NO: 20; M090 – SEQ ID NO: 21) in wild-type mice through hydrodynamic tail vein injection as described in Example 2. TNALP activity in mouse plasma was maintained to day 36 after dosing. The DNA constructs M086 (SEQ ID NO: 18) and M090 (SEQ ID NO: 21) exhibited the highest levels of plasma ALP activity, on average. FIG. 3 is an illustration of a survival graph of TNALP knockout (Akp2 (- / -)) hypophosphatasia mice, which phenotypically mimic the severe infantile form of HPP. Akp2 (- / - ) mice typically do not live longer than 20 days owing to growth failure, hypomineralization, and epileptic seizures. Mice were administered a single dose of DNA vector M090 (SEQ ID NO: 17) in an LNP carrier at a dose of 0.6 mg / kg (n=5), 1.2 mg / kg (n=8), or 1.8 mg / kg (m=8). The y-axis represents the probability of survival and the x-axis represents postnatal day (PND). FIG. 4A is a Western Blot showing comparable protein expression of TNALP in cells transfected with DNA vector M090 (no substitution), and TNALP variants in cells transfected with DNA vectors M106 (E108M), M108 (N213Q, N286Q), and M107 (E108M, N213Q, N286Q). FIG. 4B shows relative ALP activity in a 4-MUP hydrolysis assay of TNALP encoded by DNA vector M090 (no substitution), and ALP variants encoded by DNA vectors M106 (E108M), M108 (N213Q, N286Q), and M107 (E108M, N213Q, N286Q) at a DNA dose of either 100 ng / well or 200 ng / well. Average ALP activity values, expressed in relative fluorescence units per second (RFU / sec), are provided above each group data bar. DETAILED DESCRIPTION The present disclosure provides compositions and methods for the treatment of a disorder in a human patient that by using a non-integrating circular, non-viral, double stranded DNA vector having an improved expression cassette which are capable of robust and durable in vivo expression of a therapeutic protein, wherein the DNA vector is capable of forming one or more specialized secondary structures (for example, an extended cruciform structure). These improved compositions enhance the duration of transgene expression and potency of the vector, leading to improved therapy in a subject. Definitions References in the specification to “some embodiments,” “an embodiment,” “an illustrative embodiment,” etc., indicate that the embodiment described may include a particular feature, structure, or characteristic, but every embodiment may or may not necessarily include that particular feature, structure, or characteristic. Moreover, such phrases are not necessarily referring to the same embodiment. Further, when a particular feature, structure, or characteristic is described in connection with an embodiment, it is submitted that it is within the knowledge of one skilled in the art to affect such feature, structure, or characteristic in connection with other embodiments whether or not explicitly described. As used herein, the singular terms “a,” “an,” and “the” include plural referents unless context clearly indicates otherwise. Similarly, the word “or” is intended to include “and” unless the context clearly indicates otherwise. The term “includes” is defined inclusively, such that “includes A or B” means including A, B, or A and B. As used herein in the specification and in the claims, “or” should be understood to have the same meaning as “and / or” as defined above. For example, when separating items in a list, “or” or “and / or” shall be interpreted as being inclusive, e.g., the inclusion of at least about one, but also including more than one, of a number or list of elements, and, optionally, additional unlisted items. Only terms clearly indicated to the contrary, such as “only one of” or “exactly one of,” or, when used in the claims, “consisting of,” will refer to the inclusion of exactly one element of a number or list of elements. In general, the term “or” as used herein shall only be interpreted as indicating exclusive alternatives (e.g., "one or the other but not both") when preceded by terms of exclusivity, such as “either,” “one of,” “only one of,” or “exactly one of.” “Consisting essentially of,” when used in the claims, shall have its ordinary meaning as used in the field of patent law. As used herein, the terms “comprising,” “including,” “having,” and the like are used interchangeably and have the same meaning. Similarly, “comprises,” “includes,” “has,” and the like are used interchangeably and have the same meaning. Specifically, each of the terms is defined consistent with the common United States patent law definition of “comprising” and is therefore interpreted to be an open term meaning “at least about the following,” and is also interpreted not to exclude additional features, limitations, aspects, etc. Thus, for example, “a vector having components a, b, and c” means that the vector includes at least about components a, b, and c. Similarly, the phrase: “a method involving steps a, b, and c” means that the method includes at least about steps a, b, and c. Moreover, while the steps and processes may be outlined herein in a particular order, the skilled artisan will recognize that the ordering steps and processes may vary. As used herein in the specification and in the claims, the phrase “at least about one,” in reference to a list of one or more elements, should be understood to mean at least about one element selected from any one or more of the elements in the list of elements, but not necessarily including at least about one of each and every element specifically listed within the list of elements and not excluding any combinations of elements in the list of elements. This definition also allows that elements may optionally be present other than the elements specifically identified within the list of elements to which the phrase “at least about one” refers, whether related or unrelated to those elements specifically identified. Thus, as a non-limiting example, “at least about one of A and B” (or, equivalently, “at least about one of A or B,” or, equivalently “at least about one of A and / or B”) can refer, in one embodiment, to at least about one, optionally including more than one, A, with no B present (and optionally including elements other than B); in another embodiment, to at least about one, optionally including more than one, B, with no A present (and optionally including elements other than A); in yet another embodiment, to at least about one, optionally including more than one, A, and at least about one, optionally including more than one, B (and optionally including other elements); etc. As used herein, the term “about” refers to a measurable value such as an amount of the length of a nucleotide sequence, a polynucleotide or polypeptide sequence, dose, time, temperature, plasma level, transcript level, sequence identity, and the like, is meant to encompass + / - 10% of the specified amount. The term “extended cruciform” refers to a structure contained within a DNA vector formed through the interaction of two specifically oriented repeating sequences, for example AAV-derived inverted terminal repeat (ITR) sequences, separated by a small, non-repeating sequence of between about 25 and 460 base-pairs (bps), for example a non-repeating sequence comprising a small bacterial Ori, wherein the two repeating sequences are capable of aligning, forming an extended cruciform structure containing a Holliday junction and two double-stranded arms of significant length (e.g., 80-110 bps or more), with a loop formed at the end of each extended arm by the non- repeating sequence. As used herein, the term "Fc" refers to a human IgG Fc domain. Subtypes of IgG such as IgGl, IgG2, IgG3, and IgG4 are all being contemplated for usage as Fc domains. As used herein, the terms “hypophosphatasia” and “HPP” refer to a rare, heritable skeletal disorder caused by, e.g., one or more loss-of-function mutations in the ALPL (alkaline phosphatase, liver / bone / kidney) gene, which encodes tissue-nonspecific alkaline phosphatase (TNALP). HPP can be further characterized as, e.g., infantile HPP or perinatal HPP (e.g., benign perinatal HPP or lethal perinatal HPP). For instance, “infantile HPP” describes a patient having HPP that is about three years of age or younger, whereas “perinatal HPP” describes a patient having HPP immediately before or after birth (e.g., one to four weeks after birth). The age of onset of HPP, such as when the subject exhibits symptoms of HPP, can also be categorized as, e.g., perinatal-onset HPP and infantile-onset HPP. Patients with HPP can exhibit symptoms of HPP including, but not limited to, skeletal deformity, hypotonia, mobility impairments, gait disturbance, bone deformity, joint pain, bone pain, bone fracture, muscle weakness, muscle pain, rickets (e.g., defects in growth plate cartilage), premature loss of deciduous teeth, incomplete bone mineralization, elevated blood and / or urine levels of phosphoethanolamine (PEA), PPi, pyridoxal 5′-phosphate (PLP), hypomineralization, rachitic ribs, hypercalciuria, short stature, HPP-related seizure, inadequate weight gain, craniosynostosis, and / or calcium pyrophosphate dihydrate crystal deposition (CPPD) in joints leading to, e.g., chondrocalcinosis and premature death. Symptoms of HPP can also include TBM and symptoms of TBM, such as cardio-respiratory arrest, tracheostomy, cardiac arrest, respiratory distress, sputum retention, wheezing, coughing, anoxic spells, cyanosis, bradycardia, tachyarrhythmia, spontaneous hyperextension of the neck, prolonged expiratory breathing phase, failure to thrive, sternal retractions, substernal retractions, intercostal retractions, intermittent or continuous dyspnea, and recurrent bronchitis or pneumonia. As provided herein, the terms “identity”, “identical”, “homology”, “homologous”, and the like are used interchangeably to indicate similarity between two sequences. In some embodiments, a transgene sequence for insertion or incorporation into a circular, non-integrating, non-viral DNA vector having an extended cruciform structure of the present disclosure may a have a nucleic acid sequence, or encode an amino acid sequence, with at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 86%, at least about 87%, at least about 88%, at least about 89%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99.0% sequence homology or identity to a nucleic acid or amino acid sequence set forth herein. Percent sequence homology or identity is calculated by determining the number of matched positions in aligned sequences, dividing the number of matched positions by the length of an aligned sequence, and multiplying by 100. A matched position refers to a position in which identical amino acids or nucleic acids occur at the same position in aligned sequences. The percent sequence identity between a particular nucleic acid or amino acid sequence and a sequence referenced by a particular sequence homology or identification number (e.g., SEQ ID NO: 36, SEQ ID NO: 38, etc.) can be determined as follows. First, a nucleic acid or amino acid sequence is compared to the sequence set forth in a particular sequence identification number using the BLAST 2 Sequences (Bl2seq) program from the stand-alone version of BLASTZ containing BLASTN version 2.0.14 and BLASTP version 2.0.14. This stand-alone version of BLASTZ can be obtained online at fr.com / blast or at ncbi.nlm.nih.gov. Instructions explaining how to use the Bl2seq program can be found in the readme file accompanying BLASTZ. Bl2seq performs a comparison between two sequences using either the BLASTN or BLASTP algorithm. BLASTN is used to compare nucleic acid sequences, while BLASTP is used to compare amino acid sequences. To compare two nucleic acid sequences, the options are set as follows: -i is set to a file containing the first nucleic acid sequence to be compared (e.g., C:\seq1.txt); -j is set to a file containing the second nucleic acid sequence to be compared (e.g., C:\seq2.txt); -p is set to blastn; -o is set to any desired file name (e.g., C:\output.txt); -q is set to -1; -r is set to 2; and all other options are left at their default setting. For example, the following command can be used to generate an output file containing a comparison between two sequences: C:\Bl2seq -i c:\seq1.txt -j c:\seq2.txt -p blastn - o c:\output.txt -q -1 -r 2. To compare two amino acid sequences, the options of Bl2seq are set as follows: -i is set to a file containing the first amino acid sequence to be compared (e.g., C:\seq1.txt); -j is set to a file containing the second amino acid sequence to be compared (e.g., C:\seq2.txt); -p is set to blastp; -o is set to any desired file name (e.g., C:\output.txt); and all other options are left at their default setting. For example, the following command can be used to generate an output file containing a comparison between two amino acid sequences: C:\Bl2seq -i c:\seq1.txt -j c:\seq2.txt -p blastp -o c:\output.txt. If the two compared sequences share homology, then the designated output file will present those regions of homology as aligned sequences. If the two compared sequences do not share homology, then the designated output file will not present aligned sequences. Once aligned, the number of matches is determined by counting the number of positions where an identical nucleotide or amino acid residue is presented in both sequences. The percent sequence identity is determined by dividing the number of matches by the length of the sequence set forth in the identified sequence (e.g., SEQ ID NO: 36, SEQ ID NO: 38, etc.), followed by multiplying the resulting value by 100. For example, the 5' Inverted Repeat Sequence derived from AAV2 sequence of SEQ ID NO: 36 has 106 matches when aligned with the 5' Inverted Repeat Sequence derived from AAV1 sequence set forth in SEQ ID NO: 38, and is 81.5 percent identical to the sequence set forth in SEQ ID NO: 38 (i.e., 106 ÷ 130 x 100 = 81.53%). It is noted that the percent sequence identity value is rounded to the nearest tenth. For example, 81.51, 81.52, 81.53, and 81.54 is rounded down to 81.5, while 81.55, 81.56, 81.57, 81.58, and 81.59 is rounded up to 81.6. It also is noted that the length value will always be an integer. As used herein, the terms “operably linked” or “operably associated” refer to a functional relationship between two nucleic acids, wherein the expression, activity, localization, etc., of one of the sequences is controlled by, directed by, regulated by, modulated by, etc., the other nucleic acid. The two nucleic acids are said to be operably linked or operably associated or in operable association. “Operably linked” or “operably associated” can also refers to a relationship between two polypeptides wherein the expression of one of the polypeptides is controlled by, directed by, regulated by, modulated by, etc., the other polypeptide. For example, transcription of a nucleic acid is directed by an operably linked promoter; post-transcriptional processing of a nucleic acid is directed by an operably linked processing sequence; translation of a nucleic acid is directed by an operably linked translational regulatory sequence such as a translation initiation sequence; transport, stability, or localization of a nucleic acid or polypeptide is directed by an operably linked transport or localization sequence such as a secretion signal sequence; and post-translational processing of a polypeptide is directed by an operably linked processing sequence. Typically, a first nucleic acid sequence that is operably linked to a second nucleic acid sequence, or a first polypeptide that is operatively linked to a second polypeptide, is covalently linked, either directly or indirectly, to such a sequence, although any effective three-dimensional association is acceptable. One of ordinary skill in the art will appreciate that multiple nucleic acids, or multiple polypeptides, may be operably linked or associated with one another. As used herein, the term “promoter,” as used herein, refers to a DNA sequence that determines the site of transcription initiation for an RNA polymerase. Promoter sequences comprise motifs which are recognized and bound by polypeptides, i.e., transcription factors. The said transcription factors shall upon binding recruit RNA polymerases II, preferably, RNA polymerase I, II or III, more preferably, RNA polymerase II or III, and most preferably, RNA polymerase II. Thereby will be initiated the expression of a nucleic acid operatively linked to the transcription control sequence. It is to be understood that dependent on the type of nucleic acid to be expressed, expression as meant herein may comprise transcription of DNA sequences into RNA polynucleotides (as suitable for, e.g., anti-sense approaches, RNAi approaches or ribozyme approaches) or may comprise transcription of DNA sequences into RNA polynucleotides followed by translation of the said RNA polynucleotides into polypeptides (as suitable for, e.g., gene expression and recombinant polypeptide production approaches). In order to govern expression of a nucleic acid sequence, the transcription control sequence may be located immediately adjacent to the nucleic acid to be expressed, i.e., physically linked to the said nucleic acid at its 5' end. Alternatively, it may be located in physical proximity. In the latter case, however, the sequence must be located so as to allow functional interaction with the nucleic acid to be expressed. As used herein, the term “R6K replication origin” or “R6K origin of replication” or “R6K ori” refers to a sequence which is specifically recognized by the R6K Rep protein to initiate DNA replication. The term “R6K origin of replication” includes but is not limited to R6Kγ replication origin sequences disclosed herein as SEQ ID NO: 54, SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, and SEQ ID NO: 58, and variants thereof. The R6K origin of replication includes CpG free versions (e.g., SEQ ID NO: 55) as described in Droeourt et al., United States Patent No. 7244609, incorporated herein by reference. As used herein, the terms “regulatory sequence” or “regulatory element” refer to a nucleic acid sequence that regulates one or more steps in the expression (particularly transcription, but in some cases other events such as splicing or other processing) of nucleic acid sequence(s) with which it is operatively linked. The term includes promoters, enhancers, and other transcriptional control elements that direct or enhance transcription of an operatively linked nucleic acid. Regulatory sequences may direct constitutive expression (e.g., expression in most or all cell types under typical physiological conditions in culture or in an organism), cell type specific, lineage specific, or tissue specific expression, and / or regulatable (inducible or repressible) expression. For example, expression may be induced or repressed by the presence or addition of an inducing agent such as a hormone or other small molecule, by an increase in temperature, etc. Non-limiting examples of cell type, lineage, or tissue specific promoters appropriate for use in mammalian cells include lymphoid-specific promoters (see, for example, Calame et al., Adv. Immunol. 43:235, 1988) such as promoters of T cell receptors (see, e.g., Winoto et al., EMBO J. 8:729, 1989) and immunoglobulins (see, for example, Banerji et al., Cell 33:729, 1983; Queen et al., Cell 33:741, 1983), and neuron-specific promoters (e.g., the neurofilament promoter; Byrne et al., Proc. Natl. Acad. Sci. USA 86:5473, 1989). Developmentally regulated promoters include hox promoters (see, e.g., Kessel et al., Science 249:374, 1990) and the α-fetoprotein promoter (Campes et al., Genes Dev. 3:537, 1989). Some regulatory elements may inhibit or decrease expression of an operatively linked nucleic acid. Such regulatory elements may be referred to as "negative regulatory elements." A regulatory element whose activity can be induced or repressed by exposure to an inducing or repressing agent and / or by altering environmental conditions is referred to herein as a “regulatable” element. As used herein, the term “RNA selectable marker” refers to a non-translated RNA that is expressed within a plasmid or vector that regulates a chromosomally-expressed target gene in a host cell to allow for selection of the plasmid or vector from the host cell. In some embodiments, the non-translated RNA comprises a tRNA. In some embodiments, the non-translated RNA comprises an antisense repressor RNA. In some embodiments, the non-translated RNA comprises an engineered repressor RNA or a synthetic small RNA. In some embodiments, the RNA selectable marker is RNA-OUT. In some embodiments, the RNA selectable marker is RNA-IN. As used herein, the term “signal peptide” refers to a short peptide (about 5 to about 30 amino acids long) at the N-terminus of a polypeptide that directs a polypeptide towards the secretory pathway (e.g., the extracellular space). In some embodiments, the signal peptide is typically cleaved during secretion of the polypeptide. In some embodiments, the signal sequence may direct the polypeptide to an intracellular compartment or organelle. In some embodiments, a signal sequence may be identified by homology, or biological activity, to a peptide with the known function of targeting a polypeptide to a particular region of the cell. As used herein, the term “subject,” “host,” and “patient” typically refers to a human. As used herein, the terms “variants” or “variant” refer to a nucleic acid or polypeptide differing from a reference nucleic acid or polypeptide but retaining essential properties thereof. Generally, variants are overall closely similar, and, in many regions, identical to the reference nucleic acid or polypeptide. TNALP-encoding DNA Vectors with Enhanced Expression The present disclosure provides compositions, and methods of using such composition, comprising isolated, circular, non-integrating, non-viral DNA vectors having improved expression of TNALP, wherein the DNA vectors are capable of forming one or more specialized secondary structures, for example one or more extended cruciform structures, to treat a human with HPP. The DNA vectors provided herein comprise a first portion comprising a nucleic acid comprising a TNALP polypeptide expression cassette, and a second portion comprising a nucleic acid capable of forming a specialized secondary structure, such as an extended cruciform structure. The novel TNALP expression construct and specialized secondary structure in combination provide exceptional stability and reduced immunogenicity, leading to elevated TNALP transgene activity that is durably maintained for an extended length of time (see, e.g., FIG. 2), leading to increased potency for treating HPP in a subject (see, e.g., Example 2). TNALP Expression Cassette The DNA vectors provided herein comprise a first portion comprising a nucleic acid comprising a TNALP polypeptide expression cassette. The TNALP polypeptide generally comprised the formula [A]–[B]–[R]–[E]y, wherein: [A] comprises a secretion signal peptide; [B] comprises TNALP; [R] is –(M(Fc)N)–, wherein M is a dipeptide selected from -L-K-, -L-S-, or -S-S-, wherein N is the dipeptide D-I, and wherein Fc is an Fc domain; and, [E] comprises a bone targeting amino acid sequence, and y = 3-30. As provided herein, the TNALP polypeptide comprises a secretion signal peptide, which allows the TNALP to be excreted into an HPP patient’s plasma following its expression in a cell that has been transfected by a DNA vector provided herein. As provided herein, the secretion signal peptide is conjugated to the N-terminal of the TNALP polypeptide and cleaved from TNALP after translocation into the endoplasmic reticulum prior to excretion. In some embodiments, the secretion is an mIgKVIII secretion signal. In some embodiments, the secretion signal peptide comprises the amino acid sequence of SEQ ID NO: 24. In some embodiments, the secretion signal is encoded by the nucleic acid sequence of SEQ ID NO: 25, or a nucleic acid sequence at least about 80%, 85%, 90%, 95%, 97%, 98%, or 99% identical thereto. In some embodiments, the secretion signal is the wild type TNALP secretion signal. In some embodiments, the secretion signal peptide comprises the amino acid of SEQ ID NO: 60. In some embodiments, the secretion signal is an IgG2H secretion signal. In some embodiments, the secretion signal is an huIgKVIII Ig kappa light chain V-III region secretion signal. In some embodiments, the secretion signal is a huCD33tPA secretion signal, or a variant thereof. In some embodiments, the secretion signal is a Secrecon secretion signal, or a variant thereof. In some embodiments, the secretion signal is a mouse Ig kappa light chain V-III region (mIgKVIII) secretion signal, or a variant thereof. In some embodiments, the secretion signal is an alpha-lactaalbumin secretion signal. In some embodiments, the secretion signal is a native secreted alkaline phosphatase (SEAP) secretion signal. In some embodiments, the secretion signal comprises the amino acid sequence of SEQ ID NO: 74. In some embodiments, the secretion signal comprises the amino acid sequence of SEQ ID NO: 75. In some embodiments, the secretion signal comprises the amino acid sequence of SEQ ID NO: 76. In some embodiments, the secretion signal comprises the amino acid sequence of SEQ ID NO: 77. In some embodiments, the secretion signal comprises the amino acid sequence of SEQ ID NO: 78. In some embodiments, the secretion signal comprises the amino acid sequence of SEQ ID NO: 79. In some embodiments, the secretion signal comprises the amino acid sequence of SEQ ID NO: 80. In some embodiments, the secretion signal comprises the amino acid sequence of SEQ ID NO: 81. In some embodiments, the secretion signal comprises the amino acid sequence of SEQ ID NO: 82. In some embodiments, the secretion signal comprises the amino acid sequence of SEQ ID NO: 83. In some embodiments, the secretion signal is encoded by a zero CpG nucleic acid sequence. In some embodiments, the TNALP in the TNALP polypeptide is a wild type sequence. In some embodiments, the TNALP is the amino acid sequence of SEQ ID NO: 26, or an amino acid sequence at least about 90%, 95%, 97%, 98%, or 99% identical thereto. In some embodiments, the TNALP is a variant, wherein the variant TNALP polypeptide has one or more substitutions relative to the amino acid sequence of SEQ ID NO: 26. In some embodiments, the variant TNALP has at least one, at least two, at least three, at least four, at least five, or more than five amino acid substitutions selected from E108A, E108S, E108T, E108Q, E108M, E108K, E108L, E108R, E108N, E108D, E108G, E108H, E108I, E108F, E108P, E108W, E108Y, E108V, N213Q, N286Q, M384A, M384R, M384N, M384D, M384Q, M384E, M384G, M384H, M384I, M384L, M384K, M384F, M384S, M384T, M384Y, M384 V, L385V, L385K, L385A, L385N, L385H, L385S, L385T, N213Q, N286Q, N413Q, , relative to SEQ ID NO: 26 (SEQ ID NO: 85). In some embodiments, the variant TNALP has an E108X substitution, wherein X = any amino acid, relative to SEQ ID NO: 26 (SEQ ID NO: 86). In some embodiments, the variant TNALP has an E108M substation relative to SEQ ID NO: 26 (SEQ ID NO: 28). In some embodiments, the variant TNALP sequence is the amino acid sequence of SEQ ID NO: 28, or an amino acid sequence at least about 85%, 90%, 95%, 97%, 98%, or 99% identical thereto. In some embodiments, the variant TNALP has a N213Q substitution and an N286Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 87). In some embodiments, the variant TNALP has an E108M substitution, an N213Q substitution, and an N286Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 88). In some embodiments, the variant TNALP has an E108A substitution relative to SEQ ID NO: 26 (SEQ ID NO: 89). In some embodiments, the variant TNALP has an E108A substitution, an N213Q substitution, and an N286Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 90). In some embodiments, the variant TNALP has an N213Q substitution (SEQ ID NO: 95). In some embodiments, the variant TNALP has an N286Q substitution (SEQ ID NO: 96). In some embodiments, the TNALP is encoded by a zero CpG nucleic acid sequence. The TNALP polypeptides include a fragment crystallizable (Fc) immunoglobulin domain, which is fused to TNALP via a dipeptide (M) hinge. The Fc immunoglobulin domain contains the CH2 and CH3 domains of the heavy chain, along with adjoining sequences. In some embodiments, the Fc domain comprises an IgG1 Fc domain, an IgG2 Fc domain, an IgG3 Fc domain, or an IgG4 Fc domain, or a chimera thereof. In some embodiments, the Fc domain comprises an IgG1 Fc domain. In some embodiments, the Fc domain comprises a chimera of an IgG2 Fc domain and an IgG4 Fc domain. In some embodiments, the Fc domain comprises the amino acid sequence of SEQ ID NO: 30, or an amino acid sequence at least about 85%, 90%, 95%, 97%, 98%, or 99% identical thereto. In some embodiments, the nucleic acid sequence which encodes for the Fc domain is a CpG-reduced or CpG-free sequence. In some embodiments, the nucleic acid sequence which encodes for the Fc domain is the nucleic acid sequence of SEQ ID NO: 31, or a nucleic acid sequence at least about 80%, 85%, 90%, 95%, 97%, 98%, or 99% identical thereto. The TNALP polypeptide further includes a bone targeting amino acid sequence. The bone targeting amino acid sequence is fused to the Fc domain via a dipeptide hinge (N). The bone targeting amino acid sequence can be any suitable bone targeting amino acid sequence, including, for example, aspartate (D), glutamate (E), aspartate-serine-serine (DSS), glutamate-glutamate- serine (EES), and valine-histidine-histidine (VHH). In some embodiments, the bone targeting amino acid sequence [E] is aspartate. In some embodiments, the bone targeting amino acid sequence [E] is glutamate. In some embodiments, the bone targeting amino acid sequence [E] is aspartate-serine-serine (DSS). In some embodiments, the bone targeting amino acid sequence [E] is glutamate-glutamate-serine (EES). In some embodiments, the bone targeting amino acid sequence [E] is aspartate-serine-serine (DSS).In certain embodiments, the bone targeting amino acid sequence is (DSS)3-30 (SEQ ID NO: 84). In some embodiments, the bone targeting amino acid sequence is (DSS)n, wherein n = y or (E)y = (DSS)6. In some embodiments, (E)y is the amino acid sequence of SEQ ID NO: 32. In some embodiments, the (E)y is encoded by the CpG free nucleic acid sequence of SEQ ID NO: 59, or a nucleic acid at least about 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical thereto. Expression of the TNALP polypeptide is driven by a promoter nucleic acid sequence operably linked to the nucleic acid encoding the TNALP polypeptide. In some embodiment, a high-level constitutive promoter is used to drive expression of the TNALP polypeptide, such as the retroviral Rous sarcoma virus (RSN) LTR promoter (optionally with the RSV enhancer), the cytomegalovirus (CMV) promoter (optionally with the CMV enhancer) (see, e.g., Boshart et al, Cell, 41:521-530 (1985)), the dihydrofolate reductase promoter, the beta-actin promoter, the beta- active promoter linked to the enhancer derived from the cytomegalovirus (CMN) immediate early (IE) promoter, the phosphoglycerol kinase (PGK) promoter, the EFlα promoter (Invitrogen), the short elongation factor 1-alpha (EF1α-short) promoter, and the long elongation factor 1-alpha (EF1L) promoter. a viral simian virus 40 (SV40) (e.g., early or late), a Moloney murine leukemia virus (MoMLV) LTR promoter, a herpes simplex virus (HSV) (thymidine kinase) promoter, , early growth response 1 (EGR1), ferritin H (FerH), ferritin L (FerL), Glyceraldehyde 3-phosphate dehydrogenase (GAPDH), eukaryotic translation initiation factor 4A1 (EIF4A1), heat shock 70 kDa protein 5 (HSPA5), heat shock protein 90 kDa beta, member 1 (HSP90B1), heat shock protein 70 kDa (HSP70), β-kinesin ((3-KIN), the human ROSA 26 locus Orions et al., Nature Biotechnology 25, 1477-1482 (2007)), a Ubiquitin C promoter (UBC), a phosphoglycerate kinase- 1 (PGK) promoter, a cytomegalovirus enhancer / chicken β-actin (CAG) promoter, a β-actin promoter and a myeloproliferative sarcoma virus enhancer, negative control region deleted, d1587rev primer-binding site substituted (MND) promoter (Challita et al., J Virol. 69(2):748-55 (1995)). In some embodiments, the native promoter for TNALP is utilized. In some embodiments, the native promoter may be preferred when it is desired that expression of the gene should mimic the native expression. In some embodiments, the native promoter may be used when expression of the gene must be regulated temporally or developmentally, or in a tissue-specific manner, or in response to specific transcriptional stimuli. In some embodiments, other native expression control elements, such as enhancer elements, polyadenylation sites or Kozak consensus sequences may also be used to mimic the native expression. In some embodiments, the transgene product or other desirable product to be expressed is operably linked to a tissue-specific promoter. For instance, if expression in skeletal muscle is desired, a promoter active in muscle should be used. These include the promoters from genes encoding skeletal α-actin, myosin light chain 2A, dystrophin, muscle creatine kinase, as well as synthetic muscle promoters with activities higher than naturally occurring promoters (see Li et al., Nat. Biotech, 17:241-245 (1999)). Examples of promoters that are tissue-specific are known for liver [albumin, Miyatake et al. J Virol, 71:5124-32 (1997); Human thyroxine binding globulin (TBG) promoter (see Yan et al, Gene.201215;506(2):289-94 (2012), the disclosure of which is hereby incorporated by reference herein in its entirety; hepatitis B virus core promoter, Sandig et al, Gene Ther., 3:1002-9 (1996); and alpha-fetoprotein (AFP), Arbuthnot et al, Hum. Gene Ther, 7:1503-14 (1996)], bone [osteocalcin, Stein et al, Mol. Biol. Rep., 24:185-96 (1997); and bone sialoprotein, Chen et al, J Bone Miner. Res., 11:654-64 (1996)], lymphocytes (CD2, Hansal et al., J Immunol, 161:1063-8 (1998); immunoglobulin heavy chain; T cell receptor a chain), neuronal [neuron-specific enolase (NSE) promoter, Andersen et al. Cell. Mol. Neurobiol, 13:503-15 (1993); neurofilament light-chain gene, Piccioli et al., 1991, Proc. Natl. Acad. Sci. USA, 88:5611-5 (1991); and the neuron-specific vgf gene, Piccioli et al, Neuron 15:373- 84 (1995)); among others. In some embodiments, the promoter is selected from the group consisting of EF1A, EF1L (human elongation factor 1 alpha (EF1A) long), MND (a synthetic promoter that contains two elements: the U3 region of a modified Moloney murine leukemia retrovirus with long terminal repeats and an enhancer from the myeloproliferative sarcoma virus (see, e.g., Rintz et al, Prooter considerations in the design of lentiviral vectors for use in treating lysosomal storage diseases, Mol. Therapy: Methods & Clinical Development (Vol. 24), Marcy 2022, 71-87), CMV, PGK, MCU3, SFFV, CBhCD1b, CD68LPP, EFS, and UbC. In some embodiments, the promoter is a tissue specific promoter. In some embodiments, the tissue specific promoter is a liver specific promoter. In some embodiments, the liver specific promoter is a thyroxin binding globulin (TBG), an alpha 1 anti-trypsin (A1AT) promoter, a human albumin promoter (see, e.g., Miyatake et al. J Virol (1997)), a hepatitis B virus core promoter (see, e.g., Sandig et al. Gene Ther. (1996)), a TTR minimal enhancer / promoter, an alpha-antitrypsin promoter, LSP, or a variant thereof. In some embodiments, the tissue specific promoter is a muscle specific promoter (see, e.g., Skpoenkova, V.V. et al. Acta Naturae. (2021)). In some embodiments, the muscle specific promoter is a muscle creatine kinase (MCK) promoter, a muscle hybrid (MH) promoter, or a variant thereof. In some embodiments, the muscle specific promoter is a MCK promoter selected from a CK6 promoter, a chimeric MHCK7 promoter, a dMCK promoter, a tMCK promoter, a CK8 promoter, or a CK8e promoter. In some embodiments, the promoter is a constitutive promoter. In some embodiments, the constitutive promoter is selected from a cytomegalovirus (CMV) promoter, CAG promoter, a CBG promoter, a CB7 promoter, or a variant thereof. In some embodiments, the promoter is a zero CpG promoter. In some embodiments, the promoter is human elongation factor 1 alpha-long (EF1L) promoter having the nucleic acid sequence of SEQ ID NO: 34, or a nucleic acid sequence at least about 80%, 85%, 90%, 95%, 97%, 98%, or 99% identical thereto. In some embodiments, the promoter is a zero CpG EF1L promoter having the nucleic acid sequence of SEQ ID NO: 33, or a nucleic acid sequence at least about 80%, 85%, 90%, 95%, 97%, 98%, or 99% identical thereto. Transcription may be increased by inserting an enhancer sequence into the DNA vectors of the present disclosure. Enhancers are typically cis-acting elements of DNA, usually about 10 to 300 bp in length, that act on a promoter to increase its transcription. Many enhancer sequences are now known from mammalian genes (globin, elastase, albumin, alpha-fetoprotein, and insulin) and from eukaryotic cell viruses. Examples include the SV40 enhancer on the late side of the replication Ori (bp 100-270), the cytomegalovirus early promoter enhancer, the polyoma enhancer on the late side of the replication Ori, and adenovirus enhancers. The enhancer may be spliced into the vector at a position 5' or 3' to the TNALP polypeptide encoding nucleic acid sequence but is preferably located at a site 5' from the promoter. In some embodiments, the expression cassette further comprises an enhancer operably linked to the nucleic acid encoding the TNALP polypeptide. Examples of suitable enhancers include, for example, an ApoE enhancer, an alpha fetoprotein enhancer, a TTR minimal promoter / enhancer, an LSP (TH-binding globulin promoter / alpha l-microglobulin / bikunin enhancer), an alpha mic / bik enhancer, an MCK enhancer, an alpha-MHC enhancer, or a variant thereof. In some embodiments, the enhancer is an ApoE enhancer comprising the nucleic acid sequence of SEQ ID NO: 35, or a nucleic acid sequence at least about 80%, 85%, 90%, 95%, 97%, 98%, or 99% identical thereto. In some embodiments, the vectors of the present disclosure include an insulator element, e.g., a cHS insulator. In addition, 3’ nucleic acid regulatory elements operably linked to the TNALP polypeptide may further be included in the expression cassette, for example, but not limited to a transcription terminator, an RNA processing signal such as a splicing signal or a polyadenylation (poly(A)) signal, sequences that stabilize cytoplasmic mRNA, for example a Woodchuck Hepatitis Virus (WHP) post-transcriptional regulatory element (WPRE). In some embodiments, the one or more additional regulatory elements are modified to reduce or remove CpG motifs. In some embodiments, the one or more additional regulatory elements are CpG-free post-transcriptional regulatory elements such as the woodchuck post-transcriptional regulatory element (WPRE), such as the WPRE2 sequence, and / or a poly(A) polyadenylation sequence, such as the bGH2 poly(A) polyadenylation sequence. In some embodiments, the post-transcription regulatory element is a WPRE sequence comprising the nucleic acid sequence of SEQ ID NO: 61, or a nucleic acid sequence at least about 80%, 85%, 90%, 95%, 97%, 98%, or 99% identical thereto. In some embodiments, the post-transcription regulatory element is a WPRE sequence comprising the nucleic acid sequence of SEQ ID NO: 62, or a nucleic acid sequence at least about 80%, 85%, 90%, 95%, 97%, 98%, or 99% identical thereto. In some embodiments, the post-transcription regulatory element is a bGH poly(A) s sequence comprising the nucleic acid sequence of SEQ ID NO: 63, or a nucleic acid sequence at least about 80%, 85%, 90%, 95%, 97%, 98%, or 99% identical thereto. In some embodiments, the post-transcription regulatory element is a bGH poly(A) s sequence comprising the nucleic acid sequence of SEQ ID NO: 64, or a nucleic acid sequence at least about 80%, 85%, 90%, 95%, 97%, 98%, or 99% identical thereto. In some embodiments, the first portion of the DNA vector comprises: a) a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 1, wherein the nucleic acid is selected from SEQ ID NOS: 2, 3, or 4, or a nucleic acid at least about 95% identical thereto; b) a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 5, wherein the nucleic acid is SEQ ID NO: 6, or a nucleic acid at least about 95% identical thereto; or, c) a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 7, wherein the nucleic acid is SEQ ID NO: 8, or a nucleic acid at least about 95% identical thereto; d) a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 9, wherein the nucleic acid is SEQ ID NO: 10, or a nucleic acid at least about 95% identical thereto; or, e) a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 11, wherein the nucleic acid is SEQ ID NO: 12, or a nucleic acid at least about 95% identical thereto. In some embodiments, the first portion comprises a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 1. The TNALP polypeptide of SEQ ID NO: 1 comprises the following (as oriented by the N-terminus to C-terminus): [A] - a secretory signal having the amino acid sequence of SEQ ID NO: 24, as encoded by zero-CpG nucleic acid sequence of SEQ ID NO: 25; [B] – a TNALP protein having the amino acid sequence of SEQ ID NO: 26, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 27; M is the dipeptide -L-K- (hinge 1); an Fc domain having the amino acid sequence of SEQ ID NO: 30, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 31; N is the dipeptide D-I; and [E] is the bone targeting amino acid sequence -D-S-S-, wherein y=6, wherein [E] has the amino acid sequence of SEQ ID NO: 32, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 59. In some embodiments, the first portion encoding the TNALP polypeptide with the amino acid sequence of SEQ ID NO: 1 comprises a nucleic acid sequence of SEQ ID NO: 2, or a nucleic acid sequence at least about 90%, 95%, 97%, 98%, or 99% identical thereto. In addition to a nucleic acid encoding SEQ ID NO: 1, SEQ ID NO: 2 further comprises an ApoE enhancer element nucleic acid sequence of SEQ ID NO: 35, and a EF1L promoter nucleic acid sequence of SEQ ID NO: 34, wherein the ApoE enhancer element and EF1L promoter sequence are operably linked to the nucleic acid sequence encoding the TNALP polypeptide. SEQ ID NO: 2 also comprises a WPRE2 sequence (SEQ ID NO: 54), and a bGH2 poly(A) sequence (SEQ ID NO: 56). In some embodiments, the first portion encoding the TNALP polypeptide with the amino acid sequence of SEQ ID NO: 1 comprises a nucleic acid sequence of SEQ ID NO: 3, or a nucleic acid sequence at least about 90%, 95%, 97%, 98%, or 99% identical thereto. In addition to a nucleic acid encoding SEQ ID NO: 1, SEQ ID NO: 3 further comprises a zero-CpG EF1L promoter nucleic acid sequence of SEQ ID NO: 33, wherein the zero-CpG EF1L promoter sequence is operably linked to the nucleic acid sequence encoding the TNALP polypeptide. SEQ ID NO: 3 also comprises a WPRE2 sequence (SEQ ID NO: 54), and a bGH2 poly(A) sequence (SEQ ID NO: 56). In some embodiments, the first portion encoding the TNALP polypeptide with the amino acid sequence of SEQ ID NO: 1 comprises a nucleic acid sequence of SEQ ID NO: 4, or a nucleic acid sequence at least about 90%, 95%, 97%, 98%, or 99% identical thereto. In addition to a nucleic acid encoding SEQ ID NO: 1, SEQ ID NO: 4 further comprises an ApoE enhancer element nucleic acid sequence of SEQ ID NO: 35, and a zero-CpG EF1L promoter nucleic acid sequence of SEQ ID NO: 33, wherein the ApoE enhancer element and zero-CpG EF1L promoter sequence are operably linked to the nucleic acid sequence encoding the TNALP polypeptide. SEQ ID NO: 4 also comprises a WPRE2 sequence (SEQ ID NO: 54), and a bGH2 poly(A) sequence (SEQ ID NO: 56). In some embodiments, the first portion comprises a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 5. The TNALP polypeptide of SEQ ID NO: 5 comprises the following (as oriented by the N-terminus to C-terminus): [A] - a secretory signal having the amino acid sequence of SEQ ID NO: 24, as encoded by zero-CpG nucleic acid sequence of SEQ ID NO: 25; [B] – a TNALP protein having the amino acid sequence of SEQ ID NO: 26, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 27; M is the dipeptide -L-S- (hinge 2); an Fc domain having the amino acid sequence of SEQ ID NO: 30, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 31; N is the dipeptide D-I; and [E] is the bone targeting amino acid sequence -D-S-S-, wherein y=6, wherein [E] has the amino acid sequence of SEQ ID NO: 32, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 59. In some embodiments, the first portion encoding the TNALP polypeptide with the amino acid sequence of SEQ ID NO: 5 comprises a nucleic acid sequence of SEQ ID NO: 6, or a nucleic acid sequence at least about 90%, 95%, 97%, 98%, or 99% identical thereto. In addition to a nucleic acid encoding SEQ ID NO: 5, SEQ ID NO: 6 further comprises an EF1L promoter nucleic acid sequence of SEQ ID NO: 34, wherein the EF1L promoter sequence is operably linked to the nucleic acid sequence encoding the TNALP polypeptide. SEQ ID NO: 6 also comprises a WPRE2 sequence (SEQ ID NO: 54), and a bGH2 poly(A) sequence (SEQ ID NO: 56). In some embodiments, the first portion comprises a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 7. The TNALP polypeptide of SEQ ID NO: 5 comprises the following (as oriented by the N-terminus to C-terminus): [A] - a secretory signal having the amino acid sequence of SEQ ID NO: 24, as encoded by zero-CpG nucleic acid sequence of SEQ ID NO: 25; [B] – a TNALP protein having the amino acid sequence of SEQ ID NO: 26, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 27; M is the dipeptide -S-S- (hinge 3); an Fc domain having the amino acid sequence of SEQ ID NO: 30, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 31; N is the dipeptide D-I; and [E] is the bone targeting amino acid sequence -D-S-S-, wherein y=6, wherein [E] has the amino acid sequence of SEQ ID NO: 32, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 59. In some embodiments, the first portion encoding the TNALP polypeptide with the amino acid sequence of SEQ ID NO: 7 comprises a nucleic acid sequence of SEQ ID NO: 8, or a nucleic acid sequence at least about 90%, 95%, 97%, 98%, or 99% identical thereto. In addition to a nucleic acid encoding SEQ ID NO: 7, SEQ ID NO: 8 further comprises an EF1L promoter nucleic acid sequence of SEQ ID NO: 34, wherein the EF1L promoter sequence is operably linked to the nucleic acid sequence encoding the TNALP polypeptide. SEQ ID NO: 8 also comprises a WPRE2 sequence (SEQ ID NO: 54), and a bGH2 poly(A) sequence (SEQ ID NO: 56). In some embodiments, the first portion comprises a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 9. The TNALP polypeptide of SEQ ID NO: 9 comprises the following (as oriented by the N-terminus to C-terminus): [A] - a secretory signal having the amino acid sequence of SEQ ID NO: 24, as encoded by zero-CpG nucleic acid sequence of SEQ ID NO: 25; [B] – a TNALP protein having the amino acid sequence of SEQ ID NO: 28 (TNALP E108M), as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 29; M is the dipeptide -L-K- (hinge 1); an Fc domain having the amino acid sequence of SEQ ID NO: 30, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 31; N is the dipeptide D-I; and [E] is the bone targeting amino acid sequence -D-S-S-, wherein y=6, wherein [E] has the amino acid sequence of SEQ ID NO: 32, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 59. In some embodiments, the first portion encoding the TNALP polypeptide with the amino acid sequence of SEQ ID NO: 9 comprises a nucleic acid sequence of SEQ ID NO: 10, or a nucleic acid sequence at least about 90%, 95%, 97%, 98%, or 99% identical thereto. In addition to a nucleic acid encoding SEQ ID NO: 9, SEQ ID NO: 10 further comprises a zg-EF1L promoter nucleic acid sequence of SEQ ID NO: 33, wherein the EF1L promoter sequence is operably linked to the nucleic acid sequence encoding the TNALP polypeptide, as well as a WPRE2 sequence (SEQ ID NO: 54), and a bGH2 poly(A) sequence (SEQ ID NO: 56). SEQ ID NO: 10 also comprises a WPRE2 sequence (SEQ ID NO: 54), and a bGH2 poly(A) sequence (SEQ ID NO: 56). In some embodiments, the first portion comprises a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 11. The TNALP polypeptide of SEQ ID NO:1 comprises the following (as oriented by the N-terminus to C-terminus): [A] - a secretory signal having the amino acid sequence of SEQ ID NO: 24, as encoded by zero-CpG nucleic acid sequence of SEQ ID NO: 25; [B] – a TNALP protein having the amino acid sequence of SEQ ID NO: 28 (TNALP E108M), as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 29; M is the dipeptide -S-S- (hinge 3); an Fc domain having the amino acid sequence of SEQ ID NO: 30, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 31; N is the dipeptide D-I; and [E] is the bone targeting amino acid sequence -D-S-S-, wherein y=6, wherein [E] has the amino acid sequence of SEQ ID NO: 32, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 59. In some embodiments, the first portion encoding the TNALP polypeptide with the amino acid sequence of SEQ ID NO: 11 comprises a nucleic acid sequence of SEQ ID NO: 12, or a nucleic acid sequence at least about 90%, 95%, 97%, 98%, or 99% identical thereto. In addition to a nucleic acid encoding SEQ ID NO: 11, SEQ ID NO: 12 further comprises a zg-EF1L promoter nucleic acid sequence of SEQ ID NO: 33, wherein the EF1L promoter sequence is operably linked to the nucleic acid sequence encoding the TNALP polypeptide, as well as a WPRE2 sequence (SEQ ID NO: 54), and a bGH2 poly(A) sequence (SEQ ID NO: 56). SEQ ID NO: 12 also comprises a WPRE2 sequence (SEQ ID NO: 54), and a bGH2 poly(A) sequence (SEQ ID NO: 56). In an alternative aspect, the first portion of the DNA vectors described herein alternatively comprise a nucleic acid sequence encoding a TNALP polypeptide comprising the formula [A]– [B]–[R]–[E]y, wherein: [A] comprises a secretion signal peptide; [B] comprises human TNALP, wherein the TNALP is selected from a TNALP having the amino acid sequence of SEQ ID NO: 26, wherein the amino acid is encoded by a nucleic acid sequence of SEQ ID NO: 27, or a nucleic acid sequence at least about 95% identical thereto, or a human TNALP derivative having an E108M substitution, the TNALP derivative having the amino acid sequence of SEQ ID NO: 28, wherein the amino acid is encoded by a nucleic acid sequence of SEQ ID NO: 29, or a nucleic acid sequence at least about 95% identical thereto, or a human TNALP having the amino acid sequence of SEQ ID NO: 26 further comprising one or more of the following substitutions: E108X, N213X, or N286X, wherein X = any amino acid, or a human TNALP derivative having the amino acid sequence of SEQ ID NO: 85, 86, 87, 88, 89, 90, 91, 95, or 96; [R] is –(M(Fc)N)–, wherein M is the dipeptide -L-S- or -S-S-, wherein N is the dipeptide - D-I-, wherein Fc is an Fc domain having an amino acid sequence of SEQ ID NO: 30, and is encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 31, or a nucleic acid at least about 95% identical thereto; and [E] comprises a bone targeting amino acid sequence. In some embodiments, [A], a secretion signal peptide, is the amino sequence of SEQ ID NO: 24 or 60. In some embodiments, the secretion signal peptide has the amino acid sequence of SEQ ID NO: 24, and is encoded by the nucleic acid sequence of SEQ ID NO: 25, or a nucleic acid sequence at least about 95% identical thereto. In some embodiments, [A], a secretion signal peptide, is the amino sequence of SEQ ID NO: 74, 75, 76, 77, 78, 79, 80, 81, 82, or 83. In some embodiments, M is the dipeptide -L-S-. In some embodiments, M is the dipeptide -S-S-. In some embodiments, the nucleic acids of the first portion encoding the TNALP polypeptide comprise improved promoter and / or enhancer element DNA sequences, for example a zero-CpG EF1 alpha (EF1L) promoter, e.g., a promoter having the nucleic acid sequence of SEQ ID NO: 33, or a nucleic acid sequence at least 95% identical thereto, driving expression of the TNALP polypeptide. In some embodiments, an EFIL promoter having the nucleic acid sequence of SEQ ID NO: 34, or a nucleic acid at least about 95% identical thereto, drives the expression of the TNALP polypeptide. In some embodiments, the EF1L promoter is amplified by an ApoE enhancer element, e.g., an enhancer element having the nucleic acid sequence of SEQ ID NO: 35, or a nucleic acid at least 95% identical thereto. In some embodiments, the TNALP derivate comprises the amino acid of SEQ ID NO: 26, further comprising the following substitutions: E108M, N213Q, and N286Q (SEQ ID NO: 88). In some embodiments, the TNALP derivate comprises the amino acid of SEQ ID NO: 26, further comprising the following substitutions: E108A, N213Q, and N286Q (SEQ ID NO: 90). In some embodiments, the TNALP derivate comprises the amino acid of SEQ ID NO: 26, further comprising the following substitution: E108A (SEQ ID NO: 89). In some embodiments, the TNALP derivate comprises the amino acid of SEQ ID NO: 90. In some embodiments, the TNALP derivate comprises the amino acid of SEQ ID NO: 91. In some embodiments, the TNALP derivate comprises the amino acid of SEQ ID NO: 95. In some embodiments, the TNALP derivate comprises the amino acid of SEQ ID NO: 96. In addition, further regulatory elements in the TNALP expression cassette may be operatively arranged 3’ downstream of the bone-targeting peptide nucleic acid encoding sequence. For example, in some embodiments, the DNA vector comprises a woodchuck hepatitis virus post- transcriptional regulatory element 2 (WPRE2) and / or bGH2 poly(A) polyadenylation sequence. These elements further contribute to efficient transcription and expression of the transgene which lead to outstanding TNALP expression in a subject. In some embodiments, the TNALP expression cassette further comprises a WPRE regulatory element selected from a nucleic acid comprising SEQ ID NO: 61 or 62, or a nucleic acid sequence at least about 90%, at least about 95%, at least about 97%, at least about 98%, or at least about 99% identical thereto. In some embodiments, the TNALP expression cassette further comprises a bGH2 poly(A) polyadenylation sequence selected from a nucleic acid comprising SEQ ID NO: 63 or 64, or a nucleic acid sequence at least about 90%, at least about 95%, at least about 97%, at least about 98%, or at least about 99% identical thereto. In some embodiments, the first portion of the DNA vectors described herein alternatively comprise a nucleic acid sequence encoding a TNALP polypeptide comprising the formula [A]– [B]–[R]–[E]y, wherein: [A] comprises a secretion signal peptide; [B] comprises a human TNALP derivative having an E108M substitution, the TNALP derivative having the amino acid sequence of SEQ ID NO: 28, wherein the amino acid is encoded by a nucleic acid sequence of SEQ ID NO: 29, or a nucleic acid sequence at least about 95% identical thereto, or a human TNALP having the amino acid sequence of SEQ ID NO: 26 further comprising one or more of the following substitutions: E108X, N213X, or N286X, wherein X = any amino acid (SEQ ID NO: 91), or a human TNALP derivative having the amino acid sequence of SEQ ID NO: 85, 86, 87, 88, 89, 90, 91, 95, or 96; [R] is –(M(Fc)N)–, wherein M is the dipeptide -L-S- or -S-S-, wherein N is the dipeptide D-I, wherein Fc is an Fc domain having an amino acid sequence of SEQ ID NO: 30, and is encoded by the nucleic acid sequence of SEQ ID NO: 31, or a nucleic acid at least about 95% identical thereto; and, [E] comprises the bone targeting amino acid sequence -D-S-S-, wherein y = 6; wherein [E] has the amino acid sequence of SEQ ID NO: 32, as encoded by the zero-CpG nucleic acid sequence of SEQ ID NO: 59; wherein the first portion further comprises a nucleic acid sequence comprising a promoter operably linked to the nucleic acid encoding the TNALP polypeptide. In some embodiments, [A], a secretion signal peptide, is the amino sequence of SEQ ID NO: 24 or 60. In some embodiments, the secretion signal peptide has the amino acid sequence of SEQ ID NO: 24, and is encoded by the nucleic acid sequence of SEQ ID NO: 25, or a nucleic acid sequence at least about 95% identical thereto. In some embodiments, M is the dipeptide -L-K-. In some embodiments, M is the dipeptide -L-S-. In some embodiments, M is the dipeptide -S-S-. In some embodiments, the nucleic acids of the first portion encoding the TNALP polypeptide comprise improved promoter and / or enhancer element DNA sequences, for example a zero-CpG EF1L promoter, e.g., a promoter having the nucleic acid sequence of SEQ ID NO: 33, or a nucleic acid sequence at least 95% identical thereto, driving expression of the TNALP polypeptide. In some embodiments, an EFIL promoter having the nucleic acid sequence of SEQ ID NO: 34, or a nucleic acid at least about 95% identical thereto, drives the expression of the TNALP polypeptide. In some embodiments, the EF1L promoter is amplified by an ApoE enhancer element, e.g., an enhancer element having the nucleic acid sequence of SEQ ID NO: 35, or a nucleic acid at least 95% identical thereto. In some embodiments, the TNALP derivate comprises the amino acid of SEQ ID NO: 26, further comprising the following substitutions: E108M, N213Q, and N286Q (SEQ ID NO: 88). In some embodiments, the TNALP derivate comprises the amino acid of SEQ ID NO: 26, further comprising the following substitutions: E108A, N213Q, and N286Q (SEQ ID NO: 90). In some embodiments, the TNALP derivate comprises the amino acid of SEQ ID NO: 26, further comprising the following substitution: E108A (SEQ ID NO: 89). In some embodiments, the TNALP derivate comprises the amino acid of SEQ ID NO: 91. In some embodiments, the TNALP derivate comprises the amino acid of SEQ ID NO: 95. In some embodiments, the TNALP derivate comprises the amino acid of SEQ ID NO: 96. In addition, further regulatory elements in the TNALP expression cassette may be operatively arranged 3’ downstream of the bone-targeting peptide nucleic acid encoding sequence. For example, in some embodiments, the DNA vector comprises a woodchuck hepatitis virus post- transcriptional regulatory element 2 (WPRE2) and / or bGH2 poly(A) polyadenylation sequence. These elements further contribute to efficient transcription and expression of the transgene which lead to outstanding TNALP expression in a subject. In some embodiments, the TNALP expression cassette further comprises a WPRE regulatory element selected from a nucleic acid comprising SEQ ID NO: 61 or 62, or a nucleic acid sequence at least about 90%, at least about 95%, at least about 97%, at least about 98%, or at least about 99% identical thereto. In some embodiments, the TNALP expression cassette further comprises a bGH2 poly(A) polyadenylation sequence selected from a nucleic acid comprising SEQ ID NO: 63 or 64, or a nucleic acid sequence at least about 90%, at least about 95%, at least about 97%, at least about 98%, or at least about 99% identical thereto. The elements of the first portion of the DNA vector comprising a nucleic acid encoding a TNALP polypeptide described above are provided in Table 1. Table 1. TNALP Expression Cassette Elements

[0002] Inhibitory Sequences of Toll-Like Receptor 9 (TLR9) It has been well demonstrated a central role for Toll-like receptor 9 (TLR9), an immune sensor of DNA, in detecting non-viral DNA vectors and activating innate immune and CD8+ T cell responses. TLRs are a family of innate immune sensors preserved across mammalian species that are found on endosomal or plasma membranes of immune or other cells. TLR9 normally senses DNA from invasion of pathogenic DNA viruses and bacteria containing unmethylated cytosine-phosphate-guanine (CpG) motifs. After binding to TLR9, CpG-rich motifs of DNA lead to its dimerization and activates TLR9 signaling through MyD88, promoting induction of type I interferons and pro-inflammatory cytokines. Innate immune responses, such as interferon induction, trigger an antiviral state among cells, while inflammation recruits other immune cells to the site of infection and primes adaptive immune responses. One solution blocking TLR9 activation is to include specific short DNA oligonucleotides that antagonize TLR9 activation into the DNA vectors having an extended cruciform structure. In some embodiments, the vectors of the present disclosure comprise one copy of such sequence. In some embodiments, the vectors of the present disclosure comprise two or more copies of such sequence. In some embodiments, the TLR9 antagonist is the nucleic acid of SEQ ID NO: 73 (ttagggttagggttagggttagggttaggg), or a nucleic acid at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identical thereto. Specialized Secondary Structures The specialized secondary structure in the circular DNA vectors provided herein is formed through the interaction of two specifically oriented repeating sequences, for example AAV-derived inverted terminal repeat (ITR) sequences, separated by a small, non-repeating sequence, for example a non-repeating sequence comprising a small bacterial origin of replication (Ori) or other non-repeating sequence described herein. By specifically orienting these two repeating sequences to flank a non-repeating sequence of particular size, for example less than about 460 base-pairs (bps), the two repeating sequences are capable of aligning to form a specialized secondary structure, for example an extended cruciform structure containing for example a Holliday junction and two double-stranded arms of significant length—for example at least about 80 bps, at least about 90 bps, at least about 100 bps, or at least about 110 bps—with a loop formed at the end of each extended arm by the non-repeating sequence. Repeating Sequences The repeating sequences included in the second portion of the DNA vectors provided herein can be derived from any known repeating sequence capable of forming a specialized structure, for example a Holliday junction and two double-stranded arms of significant length—for example at least about 80 bps, at least about 90 bps, at least about 100 bps, or at least about 110 bps—when separated by a non-repeating sequence of sufficient length. In particular embodiments, the repeating sequences are inverted terminal repeat (ITR) sequences derived from an AAV serotype, for example, but not limited to, AAV1, AAV2, AAV3, AAV4, AAV5, AVV6, AVV7, AAV8, or AAV9. In some embodiments, each of inverted repeat sequences are nucleic acid sequences which include between about 20 to about 500 bp, from between about 20 bp and about 450 bp, from between about 20 bp to about 400 bp, from between about 20 bp to about 300 bp, from between about 20 bp to about 250 bp, from between about 20 bp to about 200 bp, from between about 20 bp to about 160 bp, from between about 60 bp to about 160 bp, from about 70 bp to about 150 bp, from about 80 bp to about 140 bp, or from about 90 bp to about 130 bp. In some embodiments, each of the inverted repeat sequences have about 115 bp, 116 bp, 117 bp, 118 bp, 119 bp, 120 bp, 121 bp, 122 bp, 123 bp, 124 bp, 125 bp, 126 bp, 127 bp, 128 bp, 129 bp, 130 bp, 131 bp, 132 bp, 133 bp, 134 bp, 135 bp, 136 bp, 137 bp, 138 bp, 139 bp, 140 bp, 141 bp, 142 bp, 143 bp, 144 bp, 145 bp, 146 bp 147 bp, 148 bp, 149 bp, or about 150 bp. In some embodiments, the ITR sequences are derived from one or more AAV serotypes, for example, but not limited to, AAV1, AAV2, AAV3, AAV4, AAV5, AVV6, AVV7, AAV8, or AAV9. Suitable AAV ITR sequences for use in the present invention include any of SEQ ID NOS: 36-53 as provided for in Table 2, or a sequence at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identical thereto. Table 2. AAV ITR Sequences

[0003] In some embodiments, each of the inverted repeat sequences has a nucleic acid sequence having at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identity to any one of SEQ ID NOS: 36 – 53. In some embodiments, each of the inverted repeat sequences have a nucleic acid sequence of any one of SEQ ID NOS: 36 – 53. In some embodiments, each of the inverted repeat sequences comprise or are derived from AAV inverted terminal repeat elements (e.g., derived from elements of an AAV serotype, such as any one of AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, and / or AAV7). For instance, each of the inverted repeat groups may include one or more of the A, A', B, B', C, C', D, and / or D' elements of inverted terminal repeat elements from one or more AAV serotypes. Particularly suitable AAV ITR elements are provided in Table 3. Table 3. ITR Element Sequences In some embodiments, the ITR used to form the extended cruciform structure comprises a 5' A element comprising at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identity to ttggccactccctctctgcgcgctdgctcgctcactgaggc (SEQ ID NO: 65). In some embodiments, the ITR used to form the extended cruciform structure comprises a 5' A element comprising ttggccactccctctctgcgcgctdgctcgctcactgaggc (SEQ ID NO: 65). In some embodiments, the ITR used to form the extended cruciform structure comprises a 3' A element comprising at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identity to gcctcagtgagcgagcgagcgcgcagagagggagtggccaa (SEQ ID NO: 66). In some embodiments, the ITR used to form the extended cruciform structure comprises a 3' A element comprising gcctcagtgagcgagcgagcgcgcagagagggagtggccaa (SEQ ID NO: 66). In some embodiments, the ITR used to form the extended cruciform structure comprising a 5' B element comprises at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identity to cgggcgacc (SEQ ID NO: 67). In some embodiments, the ITR used to form the extended cruciform structure comprises a 5' B element comprising cgggcgacc (SEQ ID NO: 67). In some embodiments, the ITR used to form the extended cruciform structure comprises a 3' B element comprising at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identity to ggtcgcccg (SEQ ID NO: 68). In some embodiments, the ITR used to form the extended cruciform structure comprises a 3' B element comprising ggtcgcccg (SEQ ID NO: 68). In some embodiments, the ITR used to form the extended cruciform structure comprises a 5' C element comprising at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identity to cgcccgggc (SEQ ID NO: 69). In some embodiments, the ITR used to form the extended cruciform structure comprises a 5' C element comprising cgcccgggc (SEQ ID NO: 69). In some embodiments, the ITR used to form the extended cruciform structure comprises a 3' C element comprising at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identity to gcccgggcg (SEQ ID NO: 70). In some embodiments, the ITR used to form the extended cruciform structure comprises a 3' C element comprising gcccgggcg (SEQ ID NO: 70). In some embodiments, the ITR used to form the extended cruciform structure comprises a 5' D element comprising at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identity to aggaacccctagtgatggag (SEQ ID NO: 71). In some embodiments, the ITR used to form the extended cruciform structure comprises a 5' D element comprising aggaacccctagtgatggag (SEQ ID NO: 71). In some embodiments, the ITR used to form the extended cruciform structure comprises a 3' D element comprising at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identity to ctccatcactaggggttcct (SEQ ID NO: 72). In some embodiments, the ITR used to form the extended cruciform structure comprises a 3' D element comprising ctccatcactaggggttcct (SEQ ID NO: 72). In some embodiments, a first inverted repeat sequence may have the Formula D–A–C–C'– B–B'–A', whereas a second inverted repeat sequence may have the Formula –A–B–B'–C–C'–A'– D'. In some embodiments, a first inverted repeat sequence may have the Formula D–A–C–C'–B– B'–A', whereas a second inverted repeat sequence may have the Formula –A–B–B'–C–C'–A'–D', but do not include a DD element as described herein. In some embodiments, a first inverted repeat sequence may have the Formula D–A–B–B'–C–C'–A', whereas a second inverted repeat sequence may have the Formula –A–C–C'–B–B'–A'–D'. In some embodiments, a first inverted repeat sequence may have the Formula D–A–B–B'–C–C'–A', whereas a second inverted repeat sequence may have the Formula –A–C–C'–B–B'–A'–D', but do not include a DD element as described herein. As provided herein, the DNA vectors capable of forming one or more specialized secondary structures as provided herein do not include or form a DD-ITR, "double D" ITR, or "DD element.” Surprisingly, Applicant has discovered that the presently disclosed non-viral DNA vectors capable of forming one or more specialized secondary structures which are devoid of a "DD element” direct higher amounts of linked heterologous gene expression and may persist in vivo as long, or longer, as those vectors which do include a "DD element." Furthermore, the structure of the repeat elements within the non-viral DNA vector of the present disclosure are not formed through the process of circularization of a viral genome (e.g., an AAV genome, or AAV vector genome) or linear DNA fragment via the inverted terminal repeat sequences. Non-repeating Nucleic Acid Sequence The repeat sequences used to form the extended arms of a specialized secondary structure are contiguous with and flank the non-repeated nucleic acid sequence. In general, the non- repeating sequence can be any suitable sequence that allows for the formation of the extended cruciform, for example a non-repeating sequence of between about 25 bps and 460 bps. The non-repeating sequence provides appropriate spacing for the ITR sequences, allowing the ITR sequences to appropriately align to form the double-stranded extended arms of the cruciform, with the non-repeating sequence forming a loop between the aligned repeating sequences at the top of each extended arm. The DNA sequence elements that lead to the formation of one or more specialized secondary structures may contain additional secondary structure that stabilizes the formation of the larger cruciform structure. Thus, in some embodiments, the non- repeating sequence may contribute additional structure to the DNA vector, thus further enhancing the stability of the DNA vector. In some embodiments, the non-repeating sequence comprises a bacterial Ori having a sequence of between about 225 bps and about 400 bps. In some embodiments, the non-repeating sequence comprises a bacterial Ori having a sequence of less than about 400 bps. In some embodiments, the non-repeating sequence comprises a bacterial Ori having a sequence of less than about 350 bps. In some embodiments, the non-repeating sequence comprises a bacterial Ori having a sequence of less than about 300 bps. In some embodiments, the non-repeating sequence comprises a bacterial Ori of between about 375 bps and about 400 bps. In some embodiments, the non-repeating sequence comprises a bacterial Ori of between about 290 bps and about 310 bps. In some embodiments, the non-repeating sequence comprises a bacterial Ori of between about 235 bps and about 260 bps. In some embodiments, the bacterial Ori is derived from pR6K (see, e.g., Rakowski et al., Plasmid R6K Replication Control. Plasmid. 2013 May; 69(3): 231–242, incorporated herein by reference). In some embodiments, the bacterial Ori is derived from R6Kγ. In some embodiments, the R6K-derived Ori has been modified to reduce CpG content, e.g., to reduce CpG content by at least about 60%, at least about 50%, at least about 40%, at least about 30%, at least about 25%, at least about 20%, at least about 10%, at least about 5%, etc. In some embodiments, the bacterial Ori includes one or more internal direct repeat sequences. In some embodiments, the non-repeating sequence comprises a bacterial Ori selected from SEQ ID NOS: 54, 55, 56, 57, or 58, or a sequence at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identical thereto. In some embodiments, the nonrepeating sequences comprises a bacterial Ori having the sequence of SEQ ID NO: 57, or a sequence at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identical thereto. In some alternative embodiments, the bacterial Ori is derived from pMB1, pBR322, ColE1, p15A, pSC101, or F1. In some embodiments, the bacterial Ori is capable of forming one or more regions of internal secondary structure which are believed to assist in helping to maintain the specialized secondary structure, for example extended cruciform structure. The nucleic acid sequences of bacterial Ori of SEQ ID NOS: 54, 55, 56, 57, or 58 are provided in Table 4. Table 4. Bacterial Origin of Replication In some embodiments, in addition to an Ori sequence, the non-repeating sequence may contain additional DNA sequences, for example, small runs of extraneous and or spacer nucleotide sequences (e.g., from about 1 bp to about 20 bps), cloning or recombination sites, and / or other sites such as LoxP sites, FRT sites, attB and attP sites or their product sites attL or attR, or alternative recombination target sites derived from these sites, e.g., Lox511 or Lox66 sites, provided that such additional DNA sequences do not unduly interfere with the formation or function of a specialized secondary structure, for example an extended cruciform. As provided herein, the DNA vectors capable of forming one or more specialized secondary structures, wherein the non-repeating sequence comprises an Ori, does not include a bacterial selection marker, for example, a drug resistance gene or an RNA-based selectable marker such as an RNA-IN or RNA- Out selectable marker, within the non-repeating sequence forming the one or more specialized secondary structures. In alternative embodiments, the non-repeating sequence comprises a heterologous gene or a portion of a heterologous gene. In alternative embodiments, the non-repeating sequence comprises a bacterial suppressor tRNA. In alternative embodiments, the non-repeating sequence comprises a bacterial RNAi repressor. In alternative embodiments, the non-repeating sequence comprises a nucleic acid sequence encoding antisense RNA. In alternative embodiments, the non- repeating sequence comprises a bacterial operator sequence. In some embodiments, the bacterial operator sequence comprises a lac operator. In some embodiments, the bacterial operator sequence comprises a tet operator. In some embodiments, the non-repeating sequence lacks a drug resistance gene. In some embodiments, the non-repeating sequence lacks a selection marker. In some embodiments, the non-repeating sequence does not include a bacterial selection marker, for example, a drug resistance gene or an RNA-based selectable marker such as an RNA-IN or RNA-Out selectable marker. Nucleic Acid Sequences Forming Specialized Secondary Structures The one or more specialized secondary structures in the circular DNA vectors provided herein are formed through the interaction of two specifically oriented repeating sequences, for example AAV-derived inverted terminal repeat (ITR) sequences, separated by a small, non- repeating sequence, for example a non-repeating sequence comprising a small bacterial Ori or other non-repeating sequence described herein. In some embodiments, the nucleic acid capable of forming the one or more specialized secondary structures, for example extended cruciform structure, comprises a first nucleic acid sequence comprising a sequence selected from SEQ ID NOS: 36, 38, 40, 42, 44, 46, 48, 50, or 52, or a nucleic acid sequence at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identical thereto; a second nucleic acid comprising a sequence selected from SEQ ID NOS: 54-58, or a nucleic acid at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identical thereto; and a third nucleic acid sequence comprising a sequence selected from SEQ ID NOS: 37, 39, 41, 43, 45, 47, 49, 51, or 53, or a nucleic acid at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identical thereto. In some embodiments, the nucleic acid capable of forming one or more specialized secondary structures, for example an extended cruciform structure, comprises a nucleic acid sequence selected from a sequence of SEQ ID NOS: 13, 14, 15, or 16 (provided in Table 5), or a nucleic acid at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identical thereto. In some embodiments, the nucleic acid sequence is SEQ ID NO: 13, or a nucleic acid at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identical thereto. In some embodiments, the nucleic acid sequence is SEQ ID NO: 14, or a nucleic acid at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identical thereto. In some embodiments, the nucleic acid sequence is SEQ ID NO: 15, or a nucleic acid at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identical thereto. In some embodiments, the nucleic acid sequence is SEQ ID NO: 16, or a nucleic acid at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identical thereto. Table 5. Second Portion Sequences

[0004] Circular, Non-integrating, Non-viral DNA Vectors having Improved Expression of TNALP In one aspect, provided herein are isolated, circular, non-integrating, non-viral DNA vectors having improved expression of TNALP, wherein the DNA vectors are capable of forming one or more specialized secondary structures, for example one or more extended cruciform structures, to treat a human with HPP. In some embodiments, the extended cruciform DNA vector has the nucleic acid sequence of SEQ ID NO: 17, or a nucleic acid sequence having at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 17 is a human TNALP (hTNALP)-encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an expression cassette comprising ApoE-EF1L-driven zg-hTNALP-zg-Fc-(DSS)6 transgene, and further comprising a woodchuck hepatitis virus post-transcriptional regulatory element 2 (WPRE2) and bGH2 poly(A) polyadenylation sequence. The first portion of the DNA Vector of SEQ ID NO: 17 comprises SEQ ID NO: 2, which encodes for the N-terminus hinge -L-K- dipeptide. The ITR-R6K Ori-ITR sequence in this vector is SEQ ID NO: 13, which forms an extended cruciform structure that provides excellent reduction in immunogenicity while also facilitating exceptional nuclear entry of the vector and outstanding gene expression exceeding previously curative levels that can be maintained for long periods of time (>280 days). In this vector, the expression cassette comprises an hTNALP-encoding transgene and a human IgG Fc domain that are devoid of CpG motifs (i.e., Zero CpG or “zg”). Fusing of the Fc domain to the TNALP increases activity of TNALP. Non-viral DNA vectors of the present disclosure having Zero CpG transgenes were shown to have even greater TNALP activity for longer periods of time with reduced immunostimulatory DNA properties, allowing for re-dosability of the vector. The expression of the transgene is driven by an EF1 alpha long (EF1L) promoter and further amplified by an apolipoprotein E (ApoE) enhancer element which increases activity of the promoter. The expression cassette also contains a bGH2 poly(A) polyadenylation sequence and a WPRE2 element that further enhances efficient transcription and expression of the transgene, respectfully. These features lead to an excellent immunogenic profile of the vector while also delivering outstanding TNALP expression in a subject. In some embodiments, the extended cruciform DNA vector has the nucleic acid sequence of SEQ ID NO: 18, or a nucleic acid sequence having at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 18 is an hTNALP-encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an expression cassette comprising a zg-EF1L-zg-driven hTNALP-zg-Fc-(DSS)6 transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The first portion of the DNA Vector of SEQ ID NO: 18 comprises SEQ ID NO: 3, which encodes for the N-terminus hinge -L-K- dipeptide. The ITR-R6K Ori-ITR sequence in this vector is SEQ ID NO: 9, which forms an extended cruciform structure that provides excellent reduction in immunogenicity and enhanced nuclear entry of the vector to facilitate robust, sustained transgene expression. In this vector, the expression cassette comprises an hTNALP-encoding transgene and a human IgG Fc domain that are Zero CpG which contributes to even greater TNALP activity for longer periods of time with reduced immunostimulatory DNA properties. The expression of the transgene is driven by an EF1 alpha long (EF1L) promoter which is also Zero CpG and further contributes to the reduced immunogenicity of the vector overall. The expression cassette also contains a bGH2 poly(A) polyadenylation sequence and a WPRE2 element that further enhances transcription and expression of the transgene, respectfully. These features lead to an excellent immunogenic profile of the vector while also delivering outstanding TNALP expression in a subject. In some embodiments, the extended cruciform DNA vector has the nucleic acid sequence of SEQ ID NO: 19, or a nucleic acid sequence having at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 19 is an hTNALP-encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an expression cassette comprising an ApoE-zg-EF1L-driven zg-hTNALP-zg-Fc-(DSS)6 transgene further comprising WPRE2 and a bGH2 poly(A) polyadenylation sequence. The first portion of the DNA Vector of SEQ ID NO: 19 comprises SEQ ID NO: 4, which encodes for the N-terminus hinge -L- K- dipeptide. The ITR-R6K Ori-ITR sequence in this vector is SEQ ID NO: 13, which forms an extended cruciform structure that provides excellent reduction in immunogenicity and enhanced nuclear entry of the vector to facilitate robust, sustained transgene expression. In this vector, the expression cassette comprises an hTNALP-encoding transgene and a human IgG Fc domain that are Zero CpG which contributes to even greater TNALP activity for longer periods of time with reduced immunostimulatory DNA properties. The expression of the transgene is driven by an EF1 alpha long (EF1L) promoter which is also Zero CpG, further contributing to the reduced immunogenicity of the vector overall, which is amplified by an ApoE enhancer element. The expression cassette also contains a bGH2 poly(A) polyadenylation sequence and a WPRE2 element that further enhances transcription and expression of the transgene, respectfully. These features lead to an excellent immunogenic profile of the vector while also delivering robust and sustained TNALP expression in a subject. In some embodiments, the extended DNA vector has the nucleic acid sequence of SEQ ID NO: 20, or a nucleic acid sequence having at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 20 is an hTNALP-encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an expression cassette comprising an EF1L-zg-hTNALP-hinge2-zg-Fc-(DSS)6 transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The first portion of the DNA Vector of SEQ ID NO: 20 comprises SEQ ID NO: 6, which encodes for the N-terminus hinge -L-S- dipeptide. The ITR-R6K Ori-ITR sequence in this vector is SEQ ID NO: 9, which forms an extended cruciform structure that provides excellent reduction in immunogenicity and enhanced nuclear entry of the vector to facilitate robust, sustained transgene expression. In this vector, the expression cassette comprises an hTNALP-encoding transgene and a human IgG Fc domain that are Zero CpG which contributes to even greater TNALP activity for longer periods of time with reduced immunostimulatory DNA properties. The hTNALP transgene and Fc are separated by a novel hinge domain (-L-S-Fc-D-I-, SEQ ID NO: 93) to further increase flexibility of the encoded polypeptide. The expression of the transgene is driven by an EF1 alpha long (EF1L) promoter. The expression cassette also contains a bGH2 poly(A) polyadenylation sequence and a WPRE2 element that further enhances transcription and expression of the transgene, respectfully. These features lead to an excellent immunogenic profile of the vector while also delivering robust and sustained TNALP expression in a subject. In some embodiments, the extended DNA vector has the nucleic acid sequence of SEQ ID NO: 21, or a nucleic acid sequence having at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identical thereto. SEQ ID NO: 21 is an hTNALP-encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an expression cassette comprising an EF1L-driven zg-hTNALP-hinge3-zg-Fc-DSS6 transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The first portion of the DNA Vector of SEQ ID NO: 21 comprises SEQ ID NO: 8, which encodes for the N-terminus hinge -S- S- dipeptide. The ITR-R6K Ori-ITR sequence in this vector is SEQ ID NO: 13, which forms an extended cruciform structure that provides excellent reduction in immunogenicity and enhanced nuclear entry of the vector to facilitate robust, sustained transgene expression. In this vector, the expression cassette comprises an hTNALP-encoding transgene and a human IgG Fc domain that are Zero CpG which contributes to even greater TNALP activity for longer periods of time with reduced immunostimulatory DNA properties. The hTNALP transgene and Fc are separated by a novel hinge domain (-S-S-Fc-D-I-, SEQ ID NO: 93) to further increase flexibility of the encoded polypeptide. The expression of the transgene is driven by an EF1 alpha long (EF1L) promoter. The expression cassette also contains a bGH2 poly(A) polyadenylation sequence and a WPRE2 element that further enhances transcription and expression of the transgene, respectfully. These features lead to an excellent immunogenic profile of the vector while also delivering robust and sustained TNALP expression in a subject. In some embodiments, the extended DNA vector has the nucleic acid sequence of SEQ ID NO: 22, or a nucleic acid sequence having at least 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 22 is an hTNALP-derivative encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an zg-EF1L- driven zg-hTNALP(E108M)-hinge1-zg-Fc-(DSS)6 transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The TNALP derivative comprises an E108M amino acid substitution in the active site of the TNALP and a secretion signal. In some embodiments, the extended DNA vector has the nucleic acid sequence of SEQ ID NO: 23, or a nucleic acid sequence having at least 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 23 is an hTNALP-derivative encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an EF1L-driven zg-hTNALP(E108M)-hinge3-zg-Fc-(DSS)6 transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The TNALP derivative comprises an E108M amino acid substitution in the active site of the TNALP and a secretion signal. In alternate embodiments, the extended DNA vector has the nucleic acid sequence of SEQ ID NO: 97, or a nucleic acid sequence having at least 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 97 is an hTNALP-derivative encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an EF1L-driven zg-hTNALP(N213Q, N286Q)-hinge3-zg-Fc-(DSS)6 transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The TNALP derivative comprises a N213Q and a N286Q amino acid substitution in the active site of the TNALP and a secretion signal. In alternate embodiments, the extended DNA vector has the nucleic acid sequence of SEQ ID NO: 98, or a nucleic acid sequence having at least 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identity thereto. SEQ ID NO: 98 is an hTNALP-derivative encoding vector comprising (i) an extended cruciform structure formed by an ITR-R6K Ori-ITR sequence, and (ii) an EF1L-driven zg-hTNALP(E108M, N213Q, N286Q)-hinge3-zg-Fc-(DSS)6 transgene further comprising a WPRE2 and bGH2 poly(A) polyadenylation sequence. The TNALP derivative comprises an E108M , a N213Q, and a N286Q amino acid substitution in the active site of the TNALP and a secretion signal. Table 6. Circular DNA Vector Sequences with Improved TNALP Expression Cassettes

[0005] In an alternative aspect, provided herein is a DNA vector having an improved expression cassette with one or more nucleic acid components that are zero-CpG, wherein the improved expression cassette has substantially fewer methylated CpG motifs compared to a DNA vector that is not zero-CpG or CpG-reduced. In some embodiments, the improved expression cassette of a DNA vector as described herein has less than about 15%, less than about 12%, less than about 10%, less than about 5%, less than about 4%, less than about 3%, less than about 2%, or less than about 1% of all CpG motifs methylated. In some embodiments, the improved expression cassette has less than about 5% of all CpG motifs methylated. In some embodiments, the improved expression cassette has less than about 4% of all CpG motifs methylated. In some embodiments, the improved expression cassette has less than about 3% of all CpG motifs methylated. In some embodiments, the improved expression cassette has less than about 2% of all CpG motifs methylated. In some embodiments, the improved expression cassette has less than about 1% of all CpG motifs methylated. In some embodiments, the improved expression cassette has less than about 125 methylated CpG motifs, less than about 100 methylated CpG motifs, less than about 75 methylated CpG motifs, less than about 50 methylated CpG motifs, less than about 25 methylated CpG motifs, or less than about 10 methylated CpG motifs per vector. In some embodiments, the improved expression cassette has less than about 150 methylated CpG motifs per vector. In some embodiments, the improved expression cassette has less than about 125 methylated CpG motifs per vector. In some embodiments, the improved expression cassette has less than about 100 methylated CpG motifs per vector. In some embodiments, the improved expression cassette has less than about 75 methylated CpG motifs per vector. In some embodiments, the improved expression cassette has less than about 50 methylated CpG motifs per vector. In some embodiments, the improved expression cassette has less than about 40 methylated CpG motifs per vector. In some embodiments, the improved expression cassette has less than about 30 methylated CpG motifs per vector. In some embodiments, the improved expression cassette has less than about 20 methylated CpG motifs per vector. In some embodiments, the improved expression cassette has less than about 10 methylated CpG motifs per vector. In some embodiments, the methylation state of an improved expression cassette of an DNA vector as described herein is determined from a sample of the DNA vector using a methylation sequencing method. In some embodiments, the methylation status of the improved expression cassette is determined by a sodium bisulfite sequencing assay, or an assay related thereto. In some embodiments, the methylation state for a sequenced fragment of the improved expression cassette nucleic acid sequence is determined based on a methylation status of each of one or more sites within the sequenced fragment, wherein a site at which the methylation status is determined is a methylation site. In some embodiments, the methylation state comprises a methylation fraction value calculated based on the methylation status of each of the one or more methylation sites within the sequenced fragment. In some embodiments, the methylation fraction value is calculated as: methylation fraction = N / M, wherein M is a total number of methylation sites located within the sequenced fragment and N is a number of methylation sites that are methylated. In some embodiments, the one or more methylation sites comprise one or more CpG dinucleotide sites. In some embodiments, the one or more methylation sites comprise one or more non-CpG dinucleotide methylation sites. Pharmaceutical Compositions In one aspect, provided herein is a pharmaceutical composition suitable for administration to a human comprising: (i) a circular, non-viral, non-integrating DNA vector as described herein; and (ii) a pharmaceutically acceptable carrier or excipient. In some embodiments, the pharmaceutical composition is formulated with a lipid-based delivery vehicle. In some embodiments, the pharmaceutical composition is formulated as lipid nanoparticles. In some embodiments, the pharmaceutical composition is formulated as lipid nanocapsules. In some embodiments, the pharmaceutical composition is formulated with one or more polymers. Another aspect of the present disclosure is directed to compositions comprising one or more circular, non-viral DNA vectors, e.g., circular, non-viral DNA vectors having a nucleic acid sequence of any one of SEQ ID NOS: 17 – 23, SEQ ID NOS: 97-98, or a nucleic acid sequence at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identical thereto. In some embodiments, the DNA vector comprises the nucleic acid sequence of SEQ ID NO: 17. In some embodiments, the DNA vector comprises the nucleic acid sequence of SEQ ID NO: 18. In some embodiments, the DNA vector comprises the nucleic acid sequence of SEQ ID NO: 19. In some embodiments, the DNA vector comprises the nucleic acid sequence of SEQ ID NO: 20. In some embodiments, the DNA vector comprises the nucleic acid sequence of SEQ ID NO: 21. In some embodiments, the DNA vector comprises the nucleic acid sequence of SEQ ID NO: 22. In some embodiments, the DNA vector comprises the nucleic acid sequence of SEQ ID NO: 23. In some embodiments, the DNA vector comprises the nucleic acid sequence of SEQ ID NO: 97. In some embodiments, the DNA vector comprises the nucleic acid sequence of SEQ ID NO: 98. In some embodiments, the present disclosure provides a composition comprising one or more of the DNA vectors capable of forming one or more specialized secondary structures as described herein and a carrier therefore (e.g., a pharmaceutically acceptable carrier). The composition desirably is a physiologically acceptable (e.g., pharmaceutically acceptable) composition, which comprises a carrier, e.g., a physiologically (e.g., pharmaceutically) acceptable carrier, and the DNA vector. Any suitable carrier can be used within the context of the present disclosure, and such carriers are well known in the art, including any of those described above. In some embodiments, the pharmaceutically acceptable carrier is a delivery vehicle, for example a lipid nanoparticle or the like. In some embodiments, the non-viral DNA vectors may be formulated with a delivery vehicle. In some embodiments, the delivery vehicle is a lipid-based delivery vehicle. In some embodiments, the delivery vehicle is a lipid nanoparticle. As used herein, the term "lipid nanoparticle" or "LNP" refers to any lipid composition that can be used to deliver a therapeutic product, including, but not limited to, liposomes or vesicles, wherein an aqueous volume is encapsulated by amphipathic lipid bilayers, or wherein the lipids coat an interior that comprises a therapeutic product, or lipid aggregates or micelles, wherein the lipid-encapsulated therapeutic product is contained within a relatively disordered lipid mixture. In some embodiments, lipid nanoparticles include lipid-based compositions with a solid lipid core stabilized by a surfactant. In some embodiments, the core lipids can be fatty acids, acyiglycerols, waxes, and mixtures of these surfactants. In some embodiments, biological membrane lipids such as phospholipids, sphingomyelins, bile salts (sodium taurocholate), and sterols (cholesterol) can be utilized as stabilizers. In some embodiments, lipid nanoparticles can be formed using defined ratios of different lipid molecules, including, but not limited to, defined ratios of one or more cationic, anionic, or neutral lipids. In some embodiments, lipid nanoparticles can encapsulate molecules, such as the disclosed non-viral DNA vectors, within an outer-membrane shell and subsequently can be contacted with target cells to deliver the encapsulated molecules (e.g., the disclosed non-viral DNA vectors) to the host cell cytosol. In some embodiments, lipid nanoparticles can be modified or functionalized with non-lipid molecules, including on their surface (e.g., CD3, CD4, CD8, CD19, CD20, CD22, CD38, CD47, CD117, transferrin, ApoE, folate, etc.). In some embodiments, lipid nanoparticles can be modified to specifically bind to one or more receptors on the surface of the target cell (e.g., 1 or more receptors, 2 or more receptors, 3 or more receptors, 4 or more receptors, etc.). By "specifically bind" is meant that the lipid nanoparticle binds to the receptors on surface of the target cell with at least about 2-fold greater affinity relative to the receptors on the surface of a non-target cell, e.g., at least about 3-fold, 4-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20- fold, 25-fold, 50-fold, or 100-fold higher. Cell surface receptors to which the modified lipid nanoparticles can bind include, but are not limited, to an integrin, transferrin receptor type 1and 2, EGF receptor, a VEGF receptor, an NGF receptor, CD3, CD4, CD7, CD8, CD19, CD20, CD22, CD33, CD43, CD38, CD56, CD69, the asialoglycoprotein receptor (ASGPR), N-acetyl-D- galactose (GalNAc) receptor, a folate receptor, and a sigma receptor. In some embodiments, the first and / or the second targeting ligand bind to the asialoglycoprotein receptor (ASGPR) or GalNAc receptor. Accordingly, in some embodiments, the modified lipid nanoparticles specifically bind to ASGPR or GalNAc receptor on surface of hepatocytes. In some embodiments, the targeting ligand used to modify the lipid nanoparticles is a carbohydrate or a carbohydrate conjugate. Carbohydrate based targeting ligands include, but are not limited to, glucose, multivalent glucose, fucose, D-mannose, multivalent mannose, lactose, multivalent lactose, D-galactose, multivalent galactose, GalNAc, multivalent GalNAc (e.g., GalNAc2 and GalNAc3), acetyl-galactosamine, N- acetyl-gulucosamine, glycosylated polyaminoacids and lectins. The term multivalent indicates that one, two, three or four monosaccharide units is present. Such monosaccharide subunits may be linked to each other through glycosidic linkages or linked to a scaffold molecule. In some embodiments, lipid nanoparticles can be single layered (unilamellar) or multi-layered (multilamellar). In some embodiments, lipid nanoparticles can be complexed with nucleic acid. Unilamellar lipid nanoparticles can be complexed with nucleic acid, wherein the nucleic acid is in the aqueous interior. In some embodiments, multilamellar lipid nanoparticles can be complexed with nucleic acid, wherein the nucleic acid is in the aqueous interior, or to form or sandwiched between. In some embodiments, liposomal particles can, for example, be formed of a mixture of zwitterionic, cationic and anionic lipids which can be saturated or unsaturated, for example 1,2- distearoyl-sn-glycero-3-phosphocholine (DSPC) (zwitterionic, saturated), 1,2-dilinoleyoxy-3- dimethylaminopropane (DlinDMA) (cationic, unsaturated), and / or 1,2-dimyristoyl-rac-glycerol (DMG) (anionic, saturated). In some embodiments, the liposomes will typically comprise helper lipids. Useful helper lipids include zwitterionic lipids, such as DPPC, DOPC, DSPC, dodecylphosphocholine, 1,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine (DOPE), and 1,2- diphytanoyl-sn-glycero-3-phosphoethanolamine (DPyPE); sterols, such as cholesterol; and PEGylated lipids, such as PEG-DMPE (PEG-conjugated 1, 2-dimyristoyl-Sn-glycero-3- phosphoethanolamine-N-[methoxy (polyethylene glycol)]) or PEG-DMG (PEG-conjugated 1,2- Dimyristoyl-sn-glycerol, methoxypolyethylene Glycol). In some embodiments, suitable PEGylated lipids include PEG2K-DMPE (PEG-conjugated 1, 2-dimyristoyl-Sn-glycero-3- phosphoethanolamine-N-[methoxy (polyethylene glycol)-2000]) or PEG2K-DMG (PEG- conjugated 1,2-Dimyristoyl-sn-glycerol, methoxypolyethylene Glycol-2000). In some embodiments, LNPs for use with the non-viral DNA vectors of the present disclosure include a zwitterionic lipid which can form liposomes, optionally in combination with at least about one cationic lipid (such as N-[1-(2,3-Dioleoyloxy)propyl]-N,N,N-trimethylammonium methyl-sulfate (DOTAPBis(2-methacryloyl)oxyethyl disulfide (DSDMA), 2,3-Dioleyloxy-1- (dimethylamino)propane (DODMA), 1,2-dilinoleyoxy-3-dimethylaminopropane (DLinDMA), N,N-dimethyl-3-aminopropane (DLenDMA), etc.). In some embodiments, the lipid nanoparticles have a mean diameter ranging from between about 20 nm to about 300 nm, e.g., from between about 20 nm to about 250 nm from between about 30 nm to about 200 nm, from between about 40 nm to about 180 nm, from between about 50 nm to about 150 nm, from between about 60 nm to about 140 nm, etc. Lipid nanoparticle particle size can be determined by quasi-elastic light scattering using, for example, a Malvern Zetasizer Nano ZS (Malvern, UK) system or electron microscope using, for example, FEI Quanta 200 Scanning Electron Microscope or FEI Tecnai Twin 120kV Transmission Electron Microscope. Examples of LNPs are described by Schoeenmaker et. al., " mRNA-lipid nanoparticle COVID-19 vaccines: Structure and stability," Int J Pharm. 2021 May 15; 601: 120586, the disclosure of which is hereby incorporated by reference herein in its entirety. Other exemplary LNPs are described by Eygeris et. al., " Chemistry of Lipid Nanoparticles for RNA Delivery," Acc Chem Res.2022 Jan 4;55(1):2-12. doi: 10.1021 / acs.accounts.1c00544. Epub 2021 Dec 1. PMID: 34850635, the disclosure of which is hereby incorporated by reference herein in its entirety. Yet other suitable LNPs for use with the non-viral DNA vectors of the present disclosure are described in United States Patent Publication Nos. 2021 / 0371877, 2022 / 0175968, 2022 / 0042035, and 2022 / 0062409, the disclosures of which are hereby incorporated by reference herein in their entireties. The non-viral DNA vectors of the present disclosure may also be formulated with one or more polymers. Various polymers or copolymers may be adapted as a vehicle for the non-viral DNA vectors of the present disclosure. Exemplary polymeric materials include poly(D,L-lactic acid-co-glycolic acid) (PLGA), poly(caprolactone) (PCL), ethylene vinyl acetate polymer (EVA), poly(lactic acid) (PLA), poly(L-lactic acid) (PLLA), poly(glycolic acid) (PGA), poly(L-lactic acid-co-glycolic acid) (PLLGA), poly(D,L-lactide) (PDLA), poly(L-lactide) (PLLA), PLGA-b- poly(ethylene glycol)-PLGA (PLGA-bPEG-PLGA), PLLA-bPEG-PLLA, PLGA-PEG-maleimide (PLGA-PEG-mal), poly(D,L-lactide-co-caprolactone), poly(D,L-lactide-co-caprolactone-co- glycolide), poly(D,L-lactide-co-PEO-co-D,L-lactide), poly(D,L-lactide-co-PPO-co-D,L-lactide), polyalkyl cyanoacralate, polyurethane, poly-L-lysine (PLL), hydroxypropyl methacrylate (HPMA), polyethyleneglycol, poly-L-glutamic acid, poly(hydroxy acids), polyanhydrides, polyorthoesters, poly(ester amides), polyamides, poly(ester ethers), polycarbonates, polyalkylenes such as polyethylene and polypropylene, polyalkylene glycols such as poly(ethylene glycol) (PEG), polyalkylene oxides (PEO), polyalkylene terephthalates such as poly(ethylene terephthalate), polyvinyl alcohols (PVA), polyvinyl ethers, polyvinyl esters such as poly(vinyl acetate), polyvinyl halides such as poly(vinyl chloride) (PVC), polyvinylpyrrolidone, polysiloxanes, polystyrene (PS), polyurethanes, derivatized celluloses such as alkyl celluloses, hydroxyalkyl celluloses, cellulose ethers, cellulose esters, nitro celluloses, hydroxypropylcellulose, carboxymethylcellulose, polymers of acrylic acids, such as poly(methyl(meth)acrylate) (PMMA), poly(ethyl(meth)acrylate), poly(butyl(meth)acrylate), poly(isobutyl(meth)acrylate), poly(hexyl(meth)acrylate), poly(isodecyl(meth)acrylate), poly(lauryl(meth)acrylate), poly(phenyl(meth)acrylate), poly(methyl acrylate), poly(isopropyl acrylate), poly(isobutyl acrylate), poly(octadecyl acrylate) (polyacrylic acids), and copolymers and mixtures thereof, polydioxanone and its copolymers, polyhydroxyalkanoates, polypropylene fumarate), polyoxymethylene, poloxamers, poly(ortho)esters, poly(butyric acid), poly(valeric acid), poly(lactide-co-caprolactone), trimethylene carbonate, polyvinylpyrrolidone, polyorthoesters, polyphosphazenes, Poly([beta]-amino esters (PBAE), and polyphosphoesters, and blends and / or block copolymers of two or more such polymers. Polymer-based systems may also include Cyclodextrin polymer (CDP)-based nanoparticles such as, for example, CDP-admantane (AD)-PEG conjugates and CDP-AD-PEG-transferrin conjugates. In some embodiments, the DNA vector is administered as a pharmaceutical composition formulated with a polymer selected from diethylaminoethyl-dextran (DEAE-D), 1,5-Dimethyl- 1,5-diazaundecamethylene polymethobromide (Polybrene), poly-DL-lactide-poly(ethylene glycol) (PELA), poly(ethylene glycol)-b-poly(L-histidine-co-L-phenylalanine) (PEGbPHF), hyaluronic acid cross-linked with PEG (HA-PEG), β-Cyclodextrin–PEI-MMP-cleavable-PEG (MMP-cleavable = GPLGIAGQC) (CDPCP), PEGylated and taxol-conjugated polymeric arginine grafted poly(disulfide amine) (APP), poly(ethylene glycol) diacrylate blended with PLL, 25 kDa poly(ethylene imine) (PEI), 2 kDa PEI cross-linked with diethylene glycol (PEI-DEG-bis-NPC), 600 Da PEI cross-linked with cyclodextrin and folic acid (PEI-CyD-FA), PEI conjugated with deoxycholic acid (PEI-DA), 1.8 kDa PEI cross-linked with cystamine (rPEI), PEI cross-linked with cystamine derivative (PCDP), PEI-b-poly(glycidyl methacrylate) (PEI-pGMA), PEI functionalized with 3-(3,4-dihydroxy-phenyl) propionic acid (catechol groups) (PEI-DPA), 1.8 kDa PEI–dibenzocyclooctyl (PEI-DBCO), PEI / hyaluronic acid, PEI / chondroitin sulfate, 2 kDa PEG grafted on 25 kDa PEI (PEG-g-PEI), PEO101–PPO56–PEO101 (Poloxamer 407), Poloxamer 407 / polycarbophil, Poloxamer PF68 and T908, Poloxamer 338 (PEO141–PPO44–PEO141) (LentiBOOST), a PAMAM-, EGFR-targeting peptide, PEG, PEGylated polyamidoamine G4, 63 kDa (PAMAM, antibody, PEG), polyphenylene 3 (PPD3), one quarter of amphiphilic polyphenylene 3 (PPD3-dendron), poly(ε-caprolactone) (PCL), PCL blended with elastin like pentapeptide (VPGVG)128 (PCL / ELP), 80 kDa PCL, Poly(lactide-co-glycolic acid) (PLGA), Poly (lactic-co-glycolic) acid and poly-L-lysine (PLL / PLGA), 75 / 25 DL-PLGA 9.4 kDa / PLL 56 kDa (PLGA / PLL), 50 / 50 (PLGA / PEG), Linear copolymer of [D-mannuronate (β1→4) L-guluronate (α1→4)]n (alginate), Alginate / poloxamer 407, Poly β-(1→4)-linked D-glucosamine (chitosan), Chitosan / β-glycerol phosphate, Poly (1→6)-linked α-D-mannose (polymannose), Cellulose- grafted poly(N,N-dimethylaminoethyl methacrylate) (Cellulose-g-P(QDMAEMA)), Poly hydroxyethyl disulfide diglycidyl ether and tobramycin (Polyaminoglycoside), β-Cyclodextrin, α- Cyclodextrin with pluronic PF68 and chondroitin sulfate or hyaluric acid (α-Cyclodextrin), Ethylene glycol diglycidyl ether (EGDE) and 3,3′-diamino-N-methyl dipropylamine (3,3′) (EDGE, 3,3’), polydopamine, catecholamines (Polynorepinephrine or polydopamine), Poly(N-(2- hydroxypropyl)methacrylamide) (pHPMA), PEG cross-linked with 1,6-hexamethylene diisocyanate and epsilon caprolactone sulfamethazine (PUSMA), Poly-arginine-g-polydisulfide amine, Copolymer of polycaprolactone diol, butyl diisocyanate, and putrescine blended PEG (Polyester urethane urea), Polystyrene coated with methyl methacrylate and divinylbenzene (polystyrene), Poly(ε-caprolactone) grafted poly(sodium styrene sulfonate) (pNaSS), Hydroxyethyl methacrylate (HEMA) with aminopropyl methacrylamide (APMA) (HEMA / APMA), Vinyl ether acrylate-functionalized poly(vinyl alcohol) (PVA-VEA), Poly(2- ethyl-2-oxazoline), Cross-linked amino ketal methacrylamide and ketal bis methacrylamide mixed with siRNA, or a DNA aptamer. In some embodiments, the DNA vector is administered as a pharmaceutical composition formulated with a natural or synthetic membrane or an implanted vehicle which improve the therapeutic activity of the DNA vector by more efficient delivery. In some embodiments, the DNA vector is formulated in an implanted delivery vehicle. Implanted delivery vehicles facilitate local administration and the enhancing of a spatiotemporal release. Methods of making and formulating implanted delivery vehicles are known to those skilled in the art (Shin, S. & Shea, L.D. Lentivirus Immobilization to Nanoparticles for Enhanced and Localized Delivery From Hydrogels. Mol Ther. 18:700-706(2010); McMahon, S.S. et al. Thermosensitive hydrogel for prolonged delivery of lentiviral vector expressing neurotrophin-3 in vitro. J Gene Med. 13(11):591-601(2011 Sep 26); Kangasniemi, L. et al. Extended release of adenovirus from silica implants in vitro and in vivo. Gene Ther.16:103-110(2009)). In some embodiments, the implanted delivery vehicle comprises a polymeric hydrogel. In some embodiments, the polymeric hydrogel is composed of hydrophilic polymers, wherein the hydrophilic polymers are selected from natural or synthetic sources. A hydrogel is an exemplary carrier for gene therapies due to their high water content and physicochemical properties that mimic the extracellular matrix (ECM) of tissues. In some embodiments, the implanted delivery vehicle is biodegradable. In some embodiments, the polymeric hydrogel comprises collagen. In some embodiments, the polymeric hydrogel comprises hyaluronic acid. In some embodiments, the polymeric hydrogel comprises gelatin. In some embodiments, the polymeric hydrogel comprises alginate hydrogels. In some embodiments, the polymeric hydrogel comprises polyethylene glycol (PEG). In some embodiments, the polymeric hydrogel comprises fibronectin. In some embodiments, the polymeric hydrogel comprises agarose. In some embodiments, the polymeric hydrogel comprises fibrin. In some embodiments, the polymeric hydrogel comprises a silica gel. In some embodiments, the implanted delivery vehicle comprises a porous or permeable membrane. Non-limiting examples of polymeric particle systems for delivery of the disclosed non- viral DNA vectors include the systems described in U.S. Pat. No. 5,543,158, U.S. Pat. No. 6,007,845, U.S. Pat. No. 6,254,890, U.S. Pat. No. 6,998,115, U.S. Pat. No. 7,727,969, U.S. Pat. No. 7,427,394, U.S. Pat. No. 8,323,698, U.S. Pat. No. 8,071,082, U.S. Pat. No. 8,105,652, US 2008 / 0268063, US 2009 / 0298710, US 2010 / 0303723, US 2011 / 0027172, US 2011 / 0065807, US 2012 / 0156135, US 2014 / 0093575, WO 2013 / 090861, the disclosures of which are hereby incorporated by reference herein in their entireties. In some embodiments, the non-viral DNA vectors may be formulated as pharmaceutically acceptable nanocapsule formulations. Nanocapsules can generally entrap compounds in a stable and reproducible way (Henry-Michelland et al., 1987; Quintanar-Guerrero et al., 1998; Douglas et al., 1987). To avoid side effects due to intracellular polymeric overloading, such ultrafine particles (sized around 0.1 μm) should be designed using polymers able to be degraded in vivo. Biodegradable polyalkyl-cyanoacrylate nanoparticles that meet these requirements are contemplated for use in the present invention. Such particles may be easily made, as described (Couvreur et al., 1980; Couvreur, 1988; zur Muhlen et al., 1998; Zambaux et al. 1998; Pinto- Alphandry et al., 1995 and U.S. Pat. No.5,145,684, specifically incorporated herein by reference in its entirety). In some embodiments, the pharmaceutical compositions including the non-viral DNA vectors of the present disclosure, and which are suitable for injectable use include sterile aqueous solutions or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions (see U.S. Pat. No.5,466,468, the disclosure of which is hereby incorporated by reference herein in its entirety). In all cases the form must be sterile and must be fluid to the extent that easy syringability exists. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms, such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (e.g., glycerol, propylene glycol, and liquid polyethylene glycol, and the like), suitable mixtures thereof, and / or vegetable oils. In some embodiments, the non-viral DNA vectors may be given to the patients with physical methods, such as electroporation, sonoporation with microbubbles, sonoporation without microbubbles, magnetofection, hydroporation, photoporation, mechanical massage, jet injection, biolistics (gene gun), hydrodynamic injection, needle injection or microinjections. Methods of Treatment The DNA vectors having improved TNALP expression cassettes as provided herein facilitate persistent and potent in vivo expression of TNALP, making them particularly suitable for use to treat subjects, including human subjects, having HPP. Without wishing to be bound by any particular theory, it is believed that the DNA vectors having improved expression cassettes that are capable of forming one or more specialized secondary structures as provided herein act like an endogenous gene in the nucleus for enhanced persistence of gene expression. In some embodiments, the DNA vectors as provided herein persist for a period of at least about 4 weeks after in vivo administration. In other embodiments, the DNA vectors as provided herein persist for a period of at least about 6 weeks after in vivo administration. In other embodiments, the DNA vectors as provided herein persist for a period of at least about 8 weeks after in vivo administration. In other embodiments, the DNA vectors as provided herein persist for a period of at least about 10 weeks after in vivo administration. In other embodiments, the DNA vectors as provided herein persist for a period of at least about 12 weeks after in vivo administration. In other embodiments, the DNA vectors as provided herein persist for a period of at least about 14 weeks after in vivo administration. In other embodiments, the DNA vectors as provided herein persist for a period of at least about 16 weeks after in vivo administration. In other embodiments, the DNA vectors as provided herein persist for a period of at least about 20 weeks after in vivo administration. In other embodiments, DNA vectors as provided herein persist for a period of at least about 32 weeks after in vivo administration. In other embodiments, the DNA vectors as provided herein persist for a period of at least about 36 weeks after in vivo administration. In other embodiments, the DNA vectors as provided herein persist for a period of at least about 48 weeks after in vivo administration. In other embodiments, the DNA vectors as provided herein persist for a period of at least about 1 year after in vivo administration. In other embodiments, the DNA vectors as provided herein persist for a period of at least about 2 years after in vivo administration. In other embodiments, the DNA vectors as provided herein persist for a period of at least about 4 years after in vivo administration. It is believed that the TNALP polypeptide transgene encoded by the expression cassette comprised by the DNA vectors as provided herein are taken up by cells and expressed at levels similar to transgene expression from viral vectors; and persist for periods similar to those of non- integrating viral vectors. Without wishing to be bound by any particular theory, it is believed that DNA vectors having an improved expression cassette as provided herein, wherein the DNA vector is capable of forming one or more specialized secondary structures, act similar to some non- integrating viral vectors by assuming molecular forms that allow for persistent gene expression. Such similarities may be related to chromatin factors that both DNA vectors provided herein and viral vectors interact with or related to chromatin structures assumed by both viral vectors and the DNA vectors of the present disclosure. Chromatin is a term that describes structures that organize DNA within the nucleus of eukaryotic cells. In its simplest conception, chromatin is composed of histone proteins that form nucleosomes on DNA. However, the spacing and modifications of these histones is complex and non-random and provide structural and signaling capacity to the protein scaffold surrounding genes or structural elements. The role of chromatin in virus biology depends largely on the type of virus, but for all viruses that transverse the nucleus, interactions with chromatin is unavoidable. The importance of chromatin dynamics in the regulation of essential viral-vector processes, including entry, gene expression, and persistence is beginning to be understood. The DNA vectors as provided herein are thought to utilize a variety of aspects of chromatin dynamics in a manner akin to viral vectors. Such attributes could include chromatin structures that promote interaction with the nuclear matrix, structures that regulate epigenetic factors influencing gene expression, structures that mediate nuclear organization, or structures that promote active transcriptionally active chromatin status among others. The present disclosure is directed to administering a therapeutically effective amount of a circular, non-integrating, non-viral DNA vector containing a specialized secondary structure, for example an extended cruciform structure, capable of expressing a TNALP polypeptide transgene or a pharmaceutical composition comprising a circular, non-integrating, non-viral DNA vector containing a specialized secondary structure, such as an extended cruciform structure, capable of expressing a TNALP polypeptide transgene to a subject having HPP. Hypophosphatasia (HPP) is a rare, heritable skeletal disease with an incidence of 1 per 100,000 births for the most severe forms of the disease. The disorder results from loss-of-function mutations in the gene encoding tissue-nonspecific alkaline phosphatase (TNALP). HPP patients present a remarkable range of symptoms, from teeth loss or osteomalacia (rickets) to almost complete absence of bone mineralization in utero. Many patients with HPP present the characteristics of skeletal deformities, short stature, muscle and bone pain, impaired mobility, and premature loss of teeth. Perinatal-onset or infantile-onset HPP can also be characterized by the presence of rachitic chest deformity, vitamin B6-dependent seizures, and failure to thrive. In particular, HPP presenting at less than six months of age is often lethal due to respiratory insufficiency, with a low survival rate at one year of age. In some embodiments, hypophosphatasia may be treated by administering a therapeutically effective amount of a pharmaceutical composition including an extended-cruciform DNA vector as described herein including one or more nucleic acid sequences encoding TNALP or a polypeptide having an amino acid sequence encoding TNALP (see, e.g., SEQ ID NOS: 17 – 23). In other embodiments, treating, mitigating, or preventing a symptom of hypophosphatasia in a mammal comprises administering a therapeutically effective amount of a pharmaceutical composition including a an extended-cruciform DNA vector including one or more nucleic acid sequences encoding TNALP or a polypeptide having an amino acid sequence encoding TNALP (see, e.g., SEQ ID NOS: 17 – 23). The methods provided herein may be carried out by administering the DNA vectors or pharmaceutical compositions by any suitable routes of administration. The route of administration can be local or systemic. Exemplary routes of administration include, for example, the nasal, pulmonary, inhalation, intraarterial, intradermal, intralesional, intramuscular, intraperitoneal, intravenous, intrathecal, intravesical, parenteral, rectal, subcutaneous, and transmucosal. In some embodiments, the non-viral DNA vectors are administered in suitably formulated pharmaceutical compositions disclosed herein either subcutaneously, intraocularly, intravitreally, parenterally, subcutaneously, intravenously, intracerebro-ventricularly, intramuscularly, intrathecally, orally, intraperitoneally, by oral or nasal inhalation, or by direct injection to one or more cells, tissues, or organs by direct injection. The methods of administration may also include those modalities as described in U.S. Pat. No.5,543,158; U.S. Pat. No. 5,641,515 and U.S. Pat. No.5,399,363, each of which are incorporated by reference herein in their entireties. In some embodiments, a subject in need of treatment is treated over a particular duration of time. In some embodiments, the duration of treatment is from about 1 week to about 10 years. In some embodiments, the duration of treatment is from about 1 week to about 5 years. In some embodiments, the duration of treatment is from about 1 week to about 1 year. In some embodiments, the duration of treatment is from about 1 week to about 6 months. In some embodiments, the duration of treatment is from about 1 week to about 3 months. In some embodiments, the duration of treatment is from about 1 week to about 1 month. In some embodiments, the duration of treatment is from about 3 months to about 5 years. In some embodiments, the duration of treatment is from about 6 months to about 5 years. In some embodiments, the duration of treatment is from about 1 year to about 5 years. Each dose may be administered over any suitable period of time. In some embodiments, the dose is administered as a bolus dose. In some embodiments, the dose is administered over a period of about 1 minute to about 4 hours. In some embodiments, the dose is administered over a period of about 1 minute to about 2 hours. In some embodiments, the dose is administered over a period of about 1 minute to about 1 hour. In some embodiments, the dose is administered over a period of about 1 minute to about 30 minutes. In some embodiments, the dose is administered over a period of about 1 minute to about 15 minutes. As noted herein, the extended-cruciform DNA vectors as described herein are amenable to redosing. In some embodiments, the circular, non-viral DNA vectors are redosed for a time period ranging from between about two weeks to about five years. In some embodiments, the pharmaceutical composition is administered according to a particular frequency. In some embodiments, the frequency is daily, every 2 days, every 3 days, every 4 days, every 5 days, every 6 days, every 7 days, every 8 days, every 9 days, every 10 days, every 11 days, every 12 days, every 13 days, or every 14 days. In some embodiments, the frequency is every 3 weeks, every 4 weeks, every 5 weeks, every 6 weeks, every 7 weeks, every 8 weeks, every 9 weeks, every 10 weeks, every 11 weeks, or every 12 weeks. In some embodiments, the frequency is every 1 month, every 2 months, every 3 months, every 4 months, every 5 months, every 6 months, every 7 months, every 8 months, every 9 months, every 10 months, every 11 months, every 12 months, every 13 months, every 14 months, every 15 months, every 16 months, every 17 months, or every 18 months. In some embodiments, the frequency is every 2 years, every 3 years, every 4 years, or every 5 years. In some embodiments, DNA vectors having improved expression cassettes as described herein have low immunogenicity, wherein the administration of the DNA vector to a host does not induce an adaptive immune response or inflammatory response in the host compared to a viral or non-viral DNA vector lacking the improved expression cassette described herein. For example, viral or non-viral DNA vectors lacking improved expression cassettes as described herein can induce about 1,000, about 2,000, or about 3,000 or more differentially expressed genes (DEGs) in a sample isolated from a host across several immune related gene ontology (GO) classifications within 2 hours and / or 24 hours following administration to the host compared to baseline levels of the genes in a sample prior to administration. GO analysis allows the identification of key biological processes that are altered in response to a given treatment condition as represented by the differentially expressed genes within the pathway. DEGs are those genes that experience either two-fold upregulation or downregulation relative to baseline levels prior to DNA vector administration. In comparison to viral or non-viral DNA vectors lacking improved expression cassettes as described herein, administration of a DNA vector having an improved expression cassette as described herein to a host induces less than about 500 DEGs, less than about 400 DEGs, less than about 300 DEGs, less than about 200 DEGs, or only about 100 or less DEGs in the host within about 2 hours and / or about 24 hours following administration compared to baseline levels prior to administration. The term “log2 fold change” as used herein represents the upregulation, or alternatively, the downregulation of a gene in a treatment condition compared to a control condition, as measured by ratios of mRNA transcript levels as determined by RNA-SEQ. This value is typically reported in logarithmic scale (base 2). Herein, “fold change” is synonymous with the expression ratio of the treatment to control condition and is represented as the log2(ratio). This is described in Quakenbush, J. Microarray data normalization and transformation. Nat Genet Suppl. 32:496- 501(2002), incorporated herein by reference. A positive log2 fold change value indicates an increase of expression, while a negative log2 fold change indicates a decrease in expression. Take, for example, wherein a two-fold increase in expression of a certain gene in treatment over control condition (ratio = 2 to 1, or 2 / 1, or 2) is observed, the log base 2 of the ratio 2 (i.e., log2(2)) equals 1. See the following series of logarithmic relationships: log2(1) = 0, log2(2) = 1, log2(1 / 2) = -1, log2(4) = 2, log2(1 / 4) = -2, etc. Although ratios are an intuitive measure of gene expression changes, ratios treat up- and down-regulated genes differently. For example, genes that are up regulated by a factor of 2 have an expression ratio of 2, whereas those that are down regulated by the same factor have an expression ratio of -0.5 (50% decrease). In order to more intuitively compare expression ratio changes, gene expression ratios are log2-transformed to produce a continuous spectrum of down- and up-regulated genes. As can be seen, the logarithms of expression ratios are treated symmetrically, so that a gene that is upregulated by a factor of 2 (i.e., 2x) compared to control (i.e., doubled) has a log2(ratio) of 1, whereas a gene that is 0.5x compared to control (i.e., halved, or a 50% decrease) has a log2(ratio) of -1. In comparison, a gene with no expression change (i.e., 1x) has a log2(ratio) equal to 0. The gene expression change descriptive term “by a factor of about 2” as used herein when referring to increased gene expression refers to a changed or experimental value that is 2-times the value of the control or baseline value (i.e., changed / experimental = 30; control / baseline = 15). When describing a decrease of expression of a gene by a “factor of greater than about 2” herein, the control or baseline value is 2-times the value of the changed or experimental value (i.e., changed / experimental = 7.5; control / baseline = 15). Therefore, log2(ratio)>1 represents an increase in gene expression by greater than a factor of 2, while 0<log2(ratio)<1 represents an increase in gene expression less than a factor of 2. For example, a gene that is upregulated by a factor of 4 compared to control (i.e., quadrupled) has a log2(ratio) of 2. As another example, administration of a non-viral DNA vector having extended cruciform structure that mediates an increase in expression of immune response gene X by a factor of 1.25 compared to control (i.e., a 25% increase in gene expression) has a log2 fold change equaling 0.322. In comparison, administration of a non-viral DNA vector without cruciform structure mediating an increase by a factor of 4 (i.e., quadrupled) in expression of the same immune response gene X compared to control (i.e., a 400% increase in gene expression) has a log2 fold change equaling 2. Similarly, a log2(ratio)<-1 represents a gene that is down regulated by more than a factor of 2 (i.e., >50% decrease), while a -1>log2(ratio)<0 represents a gene that is down regulated by less than a factor of 2 (i.e., <50% decrease). For example, a gene that is down regulated by a factor of 4 (i.e., quarter-fold change, a 75% decrease) compared to control (i.e., quartered) has a log2(ratio) of -2. As another example, administration of a non-viral DNA vector without cruciform structure which mediates a decrease in expression of innate immune response gene Y by a factor of 5 compared to control (i.e. an 80% decrease) has a log2 fold change equaling -2.32. In comparison, wherein administration of a non-viral DNA vector having extended cruciform structure mediates a decrease in expression of the same innate immune response gene Y by a factor of 1.33 compared to control (i.e., 25% decrease), the log2 fold change equals -0.42. In one aspect, provided herein is a method for the treatment of hyposphosphatasia (HPP) in a human patient comprising administering a DNA vector comprising a nucleic acid selected from SEQ ID NO: 17, 18, 19, 20, 21, 22, or 23, or a nucleic acid sequence at least about 95% identical thereto, wherein administration of the DNA vector to mice induces less than 10, less than 9, less than 8, less than 7, less than 6, or less than 5 genes that are differentially expressed in a GO immune response group selected from GO:0045087 (“innate immune response”), GO: 0002376 (“immune system process”), or GO:0006955 (immune response), or a combination thereof, at about 2-hours post-administration, at about 24-hours post-administration, to about 2-hours and 24- hours post-administration as compared to baseline levels prior to administration, whereas administration of a non-viral or viral DNA vector not having an improved expression cassette as described herein induces 10 or more, 12 or more, 14 or more, 15 or more, or 20 or more DEGs within the GO immune response groups. In some embodiments, the mice are BALB / c mice. In some embodiments, the DEGs are measured by mRNA level by next-generation sequencing or another sequencing method known to those skilled in the art. In a non-limiting exemplary assessment, the mean mRNA transcript levels of the genes is measured by RNA sequencing (RNA- SEQ) of liver samples of the mice. In some embodiments, the one or more genes analyzed in the innate immune response pathway in the mice are selected from the mouse ortholog of Ankhd1, Atg9a, C1s2, C3, Card9, Cd84, Cfh, Fes, Fga, Jchain, Klrk1, Map3k5, Masp1, Naip5, Nlrc5, Oas2, Sla2, Slamf1, Trdc, Trim11, Txk, and / or Zbtb1, or a combination thereof. In some embodiments, the one or more genes in the immune response pathway are selected from the mouse ortholog of Enpp3, Iglv1, Il6, Map3k14, Oas2, Pf4, Ppbp, Tnfrsf11b, and / or Tnfrsf22, or a combination thereof. In an additional aspect, provided herein is a method for the treatment of HPP in a human patient that minimizes transcriptional responses to the treatment in the patient, comprising administering an effective amount of a DNA vector having a nucleic acid sequence selected from SEQ ID NO: 17, 18, 19, 20, 21, 22, or 23, or a DNA vector having a nucleic acid sequence at least about 95% identical thereto, wherein administration of the DNA vector induces an expression change by about a factor of 2 or greater of mRNA transcript levels in no more than 5 expressed genes in a transcriptional-related gene ontology (GO) group selected from GO: 0045944 (positive regulation of transcription from RNA polymerase II promoter), GO: 0000122 (negative regulation of transcription from RNA polymerase II promoter), 0006397 (mRNA processing), 0008380 (RNA splicing), 0006357 (regulation of transcription from RNA polymerase II promoter), 0006355 (regulation of transcription, DNA-templated), 0006351 (transcription, DNA-templated), or a combination thereof, at about 2-hours post-administration, at about 24-hours post administration, or about 2-hours and about 24-hours post-administration compared to baseline mRNA transcript levels prior to administration. In some embodiments, the ability of the DNA vector to minimize deleterious immune responses to the treatment in a human is illustrated by administering the DNA vector to adult BALB / c mice, wherein the DNA vector induces an expression change by about a factor of 2 or greater in mean mRNA transcript levels in the mice in no more than 5 expressed genes in a transcriptional-related gene ontology (GO) group selected from mouse GO: 0045944 (positive regulation of transcription from RNA polymerase II promoter), mouse GO: 0000122 (negative regulation of transcription from RNA polymerase II promoter), mouse 0006397 (mRNA processing), mouse 0008380 (RNA splicing), mouse 0006357 (regulation of transcription from RNA polymerase II promoter), mouse 0006355 (regulation of transcription, DNA-templated), 0006351 (transcription, DNA-templated), or a combination thereof, at about 2-hours post- administration, at about 24-hours post administration, or about 2-hours and about 24-hours post- administration compared to baseline mRNA transcript levels prior to administration. In a non- limiting exemplary assessment, the transcript level of the genes is measured by RNA sequencing (RNA-SEQ) of liver samples from the mice. In some embodiments, the one or more genes analyzed in the negative regulation of transcription, DNA-templated pathway in the mice is selected from Arid4a, Bclaf1, C1d, Cir1, Cux2, Dach1, Deaf1, Foxg1, Hinfp, Id2, Ing4, Klf10, Mlxipl, Mphosph8, Nkap, Nrg1, Pura, Rest, Smyd1, Tbx2, Tmprss6, Trim 11, Trim 6, Wnt4, Zbtb20, Zfhx3, and / or Zfp263, or a combination thereof. EXAMPLES EXAMPLE 1 – PRODUCTION OF CONSTRUCTS ACCORDING TO THE PRESENT DISCLOSURE In some embodiments, a circular, non-viral DNA vector as described herein can be generated from a parental plasmid including both a marker gene and high copy number Ori (e.g., pUC or pMB1) between two loxP sites by "Cre-lox recombination." The Cre recombinase can be induced through metabolic control in the bacterial cells harboring the parental plasmid. In this instance, recombination produces two circular and supercoiled DNA molecules which are topologically unlinked, each containing a single loxP site. The one resulting circular supercoiled DNA molecule comprises the marker gene and the high copy number Ori, while the other comprises the circular non-viral DNA vector of this disclosure (FIG.1A-1C). In some embodiments, other LoxP sites such as Lox511, Lox 5171, Lox 2272, M2, M3, M7, M11, Lox 71, Lox 66, LoxPsym, or others could be used to generate the non-viral DNA vector from a parental DNA plasmid. In some embodiments, other recombinases such as PhiC31, λ integrase, and Flp recombinase could be used to generate the non-viral DNA vector. In some embodiments, recombinases such as Cre, PhiC31, λ integrase, and Flp recombinase could be produced recombinantly and applied directly to purified parental plasmid DNA to generate the non-viral DNA vector. In some embodiments, marker genes would include those the encode kanamycin resistance, spectinomycin resistance, streptomycin resistance, carbenicillin resistance, bleomycin resistance, erythromycin resistance, polymyxin B resistance, tetracycline resistance, or chloramphenicol resistance among others. In some embodiments, Ori would include those sequences from pMB1, pBR322, ColE1, p15A, pSC101, or F1. EXAMPLE 2 – ELEVATED PLASMA ALP ACTIVITY FROM HUMAN TNALP-ENCODING NON-VIRAL DNA CONSTRUCTS IN ADULT MICE Circular, non-viral DNA vectors having extended cruciform structure of ITR- R6Kγ Ori- ITR sequence of 709 bps and encoding a Zero-CpG human TNALP driven by an EF1L promoter, optionally activated using an ApoE enhancer, were analyzed for plasma activity in mice. All constructs contained woodchuck hepatitis virus post-transcriptional regulatory element 2 (WPRE2) and bGH2 poly(A) polyadenylation sequence elements. Also, the expression cassette of each vector contained sequences encoding a Zero-CpG Fc domain and the (DSS)6 (SEQ ID NO: 32) bone surface binding peptide. Two vectors, M089 (SEQ ID NO: 18) and M090 (SEQ ID NO: 21), contained novel hinge domains (-L-S-Fc-D-I- (SEQ ID NO: 92) and -S-S-Fc-D-I- (SEQ ID NO: 93), respectively) separating the hTNALP transgene and the Fc domain-encoding sequence which further increase flexibility of the encoded polypeptide. All the DNA vectors having extended cruciform structure showed persistently elevated plasma ALP activity in mouse samples collected from day 1 to day 36 after dosing (FIG. 2). The non-viral DNA vectors with the most elevated plasma ALP activity were M086 (SEQ ID NO: 18) and M090 (SEQ ID NO: 21), leading to increased potency in the subjects. EXAMPLE 3. VALIDATION OF EXPRESSION CONSTRUCTS IN AKP2- / - KNOCKOUT MURINE MODEL OF HYPOPHOSPHATASIA The effect of administration of a DNA construct having a novel hinge domain (-S-S-Fc-D- I-, SEQ ID NO: 93) as described herein on survival probability was assayed on an Akp2- / - knockout murine model of hypophosphatasia. The M090 DNA construct (SEQ ID NO: 21) formulated in an MC3-based LNP formulation or RNC-402 control were administered as a single-dose intravenous (IV) injection to Akp2- / - mice on postnatal day 2. The M090 DNA construct was administered in doses of 0.6 mg / kg, 1.2 mg / kg, and 1.8 mg / kg. Administration of RNC-402 naïve control led to a median survival of only 19 days. By comparison, all doses of M090 (SEQ ID NO: 21) formulations exhibited significantly increased survival compared to control (p<0.001), while a single dose of with M090 formulated in an LNP at 1.2 mg / kg completely rescued survival of Akp2- / - mice (FIG. 3). At a dose of 1.2 mg / kg or higher, median survival is greater than 30 days. A small subset of treated mice is being followed to assess durability. EXAMPLE 4. EVALUATION OF TNALP MUTANTS Various strategies to improve the activity of TNALP were further investigated. HEK 293T cells were transfected with DNA vectors encoding TNALP (M090 – SEQ ID NO: 19) and TNALP variants (E108M substitution (M106 – SEQ ID NO: 23), E108M, N213Q, and N286Q substitutions (M107), and N213Q and N286Q substitutions (M108)) using the FuGENE 4K, per the manufacturer’s protocol (Promega). Twenty-four hours before transfection, cells were seeded into tissue culture-treated 24-well plates (Corning) at a density of 4 x 104cells / cm2in 0.5 mL DMEM with 10% FBS (growth medium) and cultured at 37°C with 5% CO2. Following transfection of DNA vectors as described herein at 100 ng or 200 ng per well, in triplicate, cells were cultured for an additional 24 hours to allow for expression and secretion of TNALP and TNALP variants. Conditioned media (400 μL) from the HEK293T transfections were collected and stored at -20°C for 48 hours. TNALP variant activity was determined by measuring the formation of 4- methylumbelliferone as a change in fluorescence at ex / em 355 / 460 nm over time. For this assay, 100 μL of 50 mM HEPES pH 7.4, 150 mM NaCl, 1 mM MgCl2 and 10 μM 4-methylumbelliferone phosphate was added to 5 μL of 10,000X diluted conditioned media in black-walled clear bottom 96-well microtiter plates. The reactions were mixed briefly and the activity was determined by tracking the fluorescence at ex / em 355 / 460 nm over time (every 30-60s over 5-20 min) at 25°C, using a SpectraMax® iD5 (Molecular Devices) microplate reader. New DNA vectors encoding TNALP variants were generated. See Table 7 below. Table 7. DNA Construct Attributes

[0006] Activity of DNA constructs encoding TNALP or TNALP variants were analyzed for protein expression and ALP activity. Western Blotting Analysis demonstrated that all constructs expressed TNALP and TNALP variants at similar levels (FIG. 4A). In a kinetic assay of 4-MUP hydrolysis at pH 7.5, however, the M106 (SEQ ID NO: 23) construct comprising an E108M substitution exhibited a 2.5-fold increased specific activity of ALP at 200 ng / well and approximately 2.35-fold increased specific activity of ALP at 100 ng / well (FIG.4B). Although the present disclosure has been described with reference to a number of illustrative embodiments, it should be understood that numerous other modifications and embodiments can be devised by those skilled in the art that will fall within the spirit and scope of the principles of this disclosure. More particularly, reasonable variations and modifications are possible in the component parts and / or arrangements of the subject combination arrangement within the scope of the foregoing disclosure, the drawings, and the appended claims without departing from the spirit of the disclosure.

Claims

CLAIMS We claim:

1. A circular, non-viral, non-integrating DNA vector comprising: (i) a first portion comprising a nucleic acid encoding a tissue-nonspecific alkaline phosphatase (TNALP) polypeptide with the amino acid sequence of SEQ ID NO: 1, wherein the nucleic acid sequence is selected from the group consisting of SEQ ID NOS: 2, 3, 4, and a nucleic acid at least about 90% identical thereto; and, (ii) a second portion comprising a nucleic acid sequence, wherein the second portion comprises, oriented in a 5’ to 3’ direction: a) a first adeno-associated virus inverted terminal repeat (AAV-ITR) sequence; b) a non-repeating sequence comprising a bacterial origin or replication (Ori); wherein the Ori is oriented between the two AAV-ITR sequences; and, c) a second AAV-ITR sequence; wherein the Ori is between about 225 bps and about 460 bps; and, wherein the second portion lacks a bacterial selection marker.

2. The DNA vector of claim 1, wherein the first portion comprises the nucleic acid sequence of SEQ ID NO: 2, or a nucleic acid sequence at least about 90% identical thereto.

3. The DNA vector of claim 1, wherein the first portion comprises the nucleic acid sequence of SEQ ID NO: 3, or a nucleic acid sequence at least about 90% identical thereto.

4. The DNA vector of claim 1, wherein the first portion comprises the nucleic acid sequence of SEQ ID NO: 4, or a nucleic acid sequence at least about 90% identical thereto.

5. A circular, non-viral, non-integrating DNA vector comprising: (i) a first portion comprising a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 5, wherein the nucleic acid is SEQ ID NO: 6, or a nucleic acid at least about 90% identical thereto; and, (ii) a second portion comprising a nucleic acid sequence, wherein the second portion comprises: a) a first adeno-associated virus inverted terminal repeat (AAV-ITR) sequence;b) a non-repeating sequence comprising a bacterial origin or replication (Ori); wherein the Ori is oriented between the two AAV-ITR sequences; and, c) a second AAV-ITR sequence; wherein the Ori is between about 225 bps and about 460 bps; and, wherein the second portion lacks a bacterial selection marker.

6. A circular, non-viral, non-integrating DNA vector comprising: (i) a first portion comprising a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 7, wherein the amino acid sequence is encoded by nucleic acid SEQ ID NO: 8, or a nucleic acid at least about 90% identical thereto; and, (ii) a second portion comprising a nucleic acid sequence, wherein the second portion comprises, oriented in a 5’ to 3’ direction: a) a first adeno-associated virus inverted terminal repeat (AAV-ITR) sequence; b) a non-repeating sequence comprising a bacterial origin or replication (Ori); wherein the Ori is oriented between the two AAV-ITR sequences; and, c) a second AAV-ITR sequence; wherein the Ori is between about 225 bps and about 460 bps; and, wherein the second portion lacks a bacterial selection marker.

7. A circular, non-viral DNA vector comprising: (i) a first portion comprising a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 9, wherein the amino acid sequence is encoded by nucleic acid SEQ ID NO: 10, or a nucleic acid at least about 90% identical thereto. (ii) a second portion comprising a nucleic acid sequence, wherein the second portion comprises, oriented in a 5’ to 3’ direction: a) a first adeno-associated virus inverted terminal repeat (AAV-ITR) sequence; b) a non-repeating sequence comprising a bacterial origin or replication (Ori); wherein the Ori is oriented between the two AAV-ITR sequences; and, c) a second AAV-ITR sequence; wherein the Ori is between about 225 bps and about 460 bps; and, wherein the second portion lacks a bacterial selection marker.

8. A circular, non-viral DNA vector comprising: (iii) a first portion comprising a nucleic acid encoding a TNALP polypeptide with the amino acid sequence of SEQ ID NO: 11, wherein the amino acid sequence is encoded by nucleic acid SEQ ID NO: 12, or a nucleic acid at least about 90% identical thereto. (iv) a second portion comprising a nucleic acid sequence, wherein the second portion comprises, oriented in a 5’ to 3’ direction: a) a first adeno-associated virus inverted terminal repeat (AAV-ITR) sequence; b) a non-repeating sequence comprising a bacterial origin or replication (Ori); wherein the Ori is oriented between the two AAV-ITR sequences; and, c) a second AAV-ITR sequence; wherein the Ori is between about 225 bps and about 460 bps; and, wherein the second portion lacks a bacterial selection marker.

9. The DNA vector of any one of claims 1-8, wherein the first AAV-ITR is selected from the group consisting of SEQ ID NOS: 36, 38, 40, 42, 44, 46, 48, 50, 52, and a sequence at least about 90% identical thereto.

10. The DNA vector of any one of claims 1-9, wherein the first AAV-ITR is selected from the group consisting of SEQ ID NOS: 37, 39, 41, 43, 45, 47, 49, 51, 53, and a sequence at least about 90% identical thereto.

11. The DNA vector of any one of claims 1-10, wherein the Ori is selected from the group consisting of SEQ ID NOS: 54, 55, 56, 57, 58, and a sequence at least about 95% identical thereto.

12. The DNA vector of any one of claims 1-8, wherein the second portion comprises a nucleic acid sequence of SEQ ID NO: 13, or a sequence at least 95% identical thereto.

13. The DNA vector of any one of claims 1-8, wherein the second portion comprises a nucleic acid sequence of SEQ ID NO: 14, or a sequence at least 95% identical thereto.

14. The DNA vector of any one of claims 1-8, wherein the second portion comprises a nucleic acid sequence of SEQ ID NO: 15, or a sequence at least 95% identical thereto.

15. The DNA vector of any one of claims 1-8, wherein the second portion comprises a nucleic acid sequence of SEQ ID NO: 16, or a sequence at least 95% identical thereto.

16. The DNA vector of any one of claims 1-15, wherein the first portion further comprises a post-transcriptional regulatory element.

17. The DNA vector of claim 16, wherein the post-transcriptional regulatory element is a woodchuck post-transcriptional regulatory element (WPRE).

18. The DNA vector of claim 17, wherein the WPRE is selected from the group consisting of SEQ ID NO: 61 and SEQ ID NO:

62.

19. The DNA vector of any one of claims 1-18, wherein the first portion further comprises a polyadenylation (poly(A)) sequence.

20. The DNA vector of claim 19, wherein the poly(A) sequence is a bovine growth hormone poly(A) sequence selected from the group consisiting of SEQ ID NO: 63 and SEQ ID NO:

64.

21. A circular, non-viral, non-integrating DNA vector comprising a nucleic acid sequence of SEQ ID NO: 17, or a nucleic acid sequence at least about 90% identical thereto.

22. A circular, non-viral, non-integrating DNA vector comprising a nucleic acid sequence of SEQ ID NO: 18, or a nucleic acid sequence at least about 90% identical thereto.

23. A circular, non-viral, non-integrating DNA vector comprising a nucleic acid sequence of SEQ ID NO: 19, or a nucleic acid sequence at least about 90% identical thereto.

24. A circular, non-viral, non-integrating DNA vector comprising a nucleic acid sequence of SEQ ID NO: 20, or a nucleic acid sequence at least about 90% identical thereto.

25. A circular, non-viral, non-integrating DNA vector comprising a nucleic acid sequence of SEQ ID NO: 21, or a nucleic acid sequence at least about 90% identical thereto.

26. A circular, non-viral, nonintegrating DNA vector comprising a nucleic acid sequence of SEQ ID NO: 22, or a nucleic acid sequence at least about 90% identical thereto.

27. A circular, non-viral, nonintegrating DNA vector comprising a nucleic acid sequence of SEQ ID NO: 23, or a nucleic acid sequence at least about 90% identical thereto.

28. A circular, non-viral, nonintegrating DNA vector comprising a nucleic acid sequence of SEQ ID NO: 97, or a nucleic acid sequence at least about 90% identical thereto.

29. A circular, non-viral, nonintegrating DNA vector comprising a nucleic acid sequence of SEQ ID NO: 98, or a nucleic acid sequence at least about 90% identical thereto.

30. A circular, non-viral, non-integrating DNA vector comprising:(i) a first portion comprising a nucleic acid encoding a TNALP polypeptide comprising the formula [A]–[B]–[R]–[E]y, wherein: [A] comprises a secretion signal peptide; [B] comprises TNALP; [R] is –(M(Fc)N)–, wherein M is the dipeptide -L-S- or -S-S-, wherein Fc is an Fc domain, and wherein N is the dipeptide D-I; [E] comprises a bone targeting amino acid sequence, wherein y = 3-30; wherein the first portion further comprises a nucleic acid sequence comprising a promoter operably linked to the nucleic acid encoding the TNALP polypeptide; and, (ii) a second portion comprising a nucleic acid sequence, wherein the second portion comprises, oriented in a 5’ to 3’ direction: a) a first adeno-associated virus inverted terminal repeat (AAV-ITR) sequence; b) a non-repeating sequence comprising a bacterial origin or replication (Ori); wherein the Ori is oriented between the two AAV-ITR sequences; and, c) a second AAV-ITR sequence; ; wherein the Ori is between about 225 bps and about 460 bps; and, wherein the second portion lacks a bacterial selection marker.

31. The DNA vector of claim 30, wherein [A] comprises a secretion signal peptide selected from an amino acid sequence of SEQ ID NOS: 24, 60, or 74-83.

32. The DNA vector of claim 31, wherein the signal peptide comprises the amino acid of SEQ ID NO: 24, and wherein the amino acid is encoded by the nucleic acid sequence of SEQ ID NO: 25, or a nucleic acid sequence at least about 90% identical thereto.

33. The DNA vector of any one of claims 30-32, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence is encoded by a nucleic acid sequence of SEQ ID NO: 27, or a nucleic acid sequence at least about 90% identical thereto.

34. The DNA vector of any one of claims 30-32, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises at least one amino acid substitutions selected from thegroup consisting of E108A, E108S, E108T, E108Q, E108M, E108K, E108L, E108R, E108N, E108D, E108G, E108H, E108I, E108F, E108P, E108W, E108Y, E108V, N213Q, N286Q, M384A, M384R, M384N, M384D, M384Q, M384E, M384G, M384H, M384I, M384L, M384K, M384F, M384S, M384T, M384Y, M384 V, L385V, L385K, L385A, L385N, L385H, L385S, L385T, N213Q, N286Q, and N413Q, relative to SEQ ID NO: 26 (SEQ ID NO: 85).

35. The DNA vector of any one of claims 30-32, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises an E108X substitution, wherein X = any amino acid relative to SEQ ID NO: 26 (SEQ ID NO: 86).

36. The DNA vector of any one of claims 30-32, wherein [B] comprises TNAPLP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises a N213Q substitution and a N286Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 87).

37. The DNA vector of any one of claims 30-32, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises an E108M substitution, an N213Q substitution, and an N286Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 88).

38. The DNA vector of any one of claims 30-32, wherein [B] comprises TNAPLP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises an E108A substitution relative to SEQ ID NO: 26 (SEQ ID NO: 89).

39. The DNA vector of any one of claims 30-32, wherein [B] comprises TNAPLP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises an E108A substitution, a N213Q substitution, and a N286Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 90).

40. The DNA vector of any one of claims 30-32, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises an E108X substitution, a N213X substitution, and a N286X substitution, wherein X = any amino acid relative to SEQ ID NO: 26 (SEQ ID NO: 91).

41. The DNA vector of any one of claims 28-30, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises a N213Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 95).

42. The DNA vector of any one of claims 30-32, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises an E108X substitution, a N286Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 96).

43. The DNA vector of any one of claims 30-42, wherein the Fc domain comprises an amino acid sequence of SEQ ID NO: 30, wherein the amino acid sequence is encoded by the nucleic acid sequence of SEQ ID NO: 31, or a nucleic acid sequence at least about 90% identical thereto.

44. The DNA vector of any one of claims 30-43, wherein [R] comprises an amino acid sequence of SEQ ID NO: 92, or an amino acid sequence at least about 90% identical thereto.

45. The DNA vector of any one of claims 30-43, wherein [R] comprises an amino acid sequence of SEQ ID NO: 93, or an amino acid sequence at least about 90% identical thereto.

46. The DNA vector of any one of claims 30-45, wherein [E] is the amino acid sequence DSS.

47. The DNA vector of claim 46, wherein y = 6, wherein [E]y comprises an amino acid sequence of SEQ ID NO: 32, wherein the amino acid sequence is encoded by the nucleic acid sequence of SEQ ID NO: 59, or a nucleic acid sequence at least about 90% identical thereto.

48. The DNA vector of any one of claims 30-45, wherein [E] is selected from aspartate, glutamate, glutamate-glutamate-serine (EES), or valine-histidine-histidine (VHH).

49. The DNA vector of any one of claims 30-48, wherein the promoter is an EF1L promoter.

50. The DNA vector of any one of claims 30-48, wherein the promoter comprises a nucleic acid sequence of SEQ ID NO: 33, or an amino acid sequence at least about 90% identical thereto.

51. The DNA vector of any one of claims 30-48, wherein the promoter comprises a nucleic acid sequence of SEQ ID NO: 34, or an amino acid sequence at least about 90% identical thereto.

52. The DNA vector of any one of claims 30-51, wherein the first portion further comprises an enhancer nucleic acid sequence.

53. The DNA vector of claim 52, wherein the enhancer nucleic acid sequence comprises SEQ ID NO: 35, or a nucleic acid sequence at least about 90% identical thereto.

54. The DNA vector of any one of claims 30-53, wherein the first portion further comprises a post-transcriptional regulatory element.

55. The DNA vector of claim 54, wherein the post-transcriptional regulatory element is a woodchuck post-transcriptional regulatory element (WPRE).

56. The DNA vector of claim 55, wherein the WPRE is selected from the group consisting of SEQ ID NOS: 61,SEQ ID NO: 62, and a nucleic acid sequence at least about 90% identical thereto.

57. The DNA vector of any one of claims 30-56, wherein the first portion further comprises a polyadenylation (poly(A)) sequence.

58. The DNA vector of claim 57, wherein the poly(A) sequence is a bovine growth hormone poly(A) sequence selected from the group consisiting of SEQ ID NO: 63, SEQ ID NO: 64, and a nucleic acid sequence at least about 90% identical thereto.

59. The DNA vector of any one of claims 30-58, wherein the first AAV-ITR is selected from the group consisting of SEQ ID NOS: 36, 38, 40, 42, 44, 46, 48, 50, 52, and a sequence at least about 90% identical thereto.

60. The DNA vector of any one of claims 30-59, wherein the first AAV-ITR is selected from the group consisting of SEQ ID NOS: 37, 39, 41, 43, 45, 47, 49, 51, 53, and a sequence at least about 90% identical thereto.

61. The DNA vector of any one of claims 30-60, wherein the Ori is selected from the group consisting of SEQ ID NOS: 54, 55, 56, 57, 58, and a sequence at least about 90% identical to SEQ ID NO: 54, a sequence at least about 90% identical to SEQ ID NO: 55, a sequence at least about 90% identical to SEQ ID NO: 56, a sequence at least about 90% identical to SEQ ID NO: 57, and a sequence at least about 90% identical to SEQ ID NO: 58.

62. The DNA vector of any one of claims 30-61, wherein the second portion comprises a nucleic acid sequence of SEQ ID NO: 13, or a sequence at least 90% identical thereto.

63. The DNA vector of any one of claims 30-61, wherein the second portion comprises a nucleic acid sequence of SEQ ID NO: 14, or a sequence at least 90% identical thereto.

64. The DNA vector of any one of claims 30-61, wherein the second portion comprises a nucleic acid sequence of SEQ ID NO: 15, or a sequence at least 90% identical thereto.

65. The DNA vector of any one of claims 30-61, wherein the second portion comprises a nucleic acid sequence of SEQ ID NO: 16, or a sequence at least 90% identical thereto.

66. A circular, non-viral, non-integrating DNA vector comprising: (i) a first portion comprising a nucleic acid encoding a TNALP polypeptide comprising the formula [A]–[B]–[R]–[E]y, wherein: [A] comprises a secretion signal peptide; [B] comprises human TNALP, wherein the TNALP is selected from a human TNALP derivative having an E108M substitution, the TNALP derivative having the amino acid sequence of SEQ ID NO: 28, wherein the amino acid is encoded by a nucleic acid sequence of SEQ ID NO: 29, or a nucleic acid sequence at least about 90% identical thereto, or a human TNALP derivative having the amino acid sequence of SEQ ID NO: 26, and further comprising one or more of the following substitutions: E108X, N213X, or N286X, wherein X = any amino acid (SEQ ID NO: 91), or a human TNALP derivative having the amino acid sequence of SEQ ID NO: 85, 86, 87, 88, 89, 90, 91, 95, or 96 [R] is –(M(Fc)N)–, wherein M is the dipeptide -L-K-, -L-S- or -S-S-, wherein Fc is an Fc domain, and wherein N is the dipeptide D-I; [E] comprises a bone targeting amino acid sequence, wherein y = 3-30; wherein the first portion further comprises a nucleic acid sequence comprising a promoter operably linked to the nucleic acid encoding the TNALP polypeptide; and, (ii) a second portion comprising a nucleic acid sequence, wherein the second portion comprises, oriented in a 5’ to 3’ direction: a first adeno-associated virus inverted terminal repeat (AAV-ITR) sequence; a non-repeating sequence comprising a bacterial origin or replication (Ori);wherein the Ori is oriented between the two AAV-ITR sequences; and, a second AAV-ITR sequence; wherein the Ori is between about 225 bps and about 460 bps; and, wherein the second portion lacks a bacterial selection marker.

67. The DNA vector of claim 66, wherein [A] comprises a secretion signal peptide selected from of the group consisting of SEQ ID NOS: 24, 60, and 74-83.

68. The DNA vector of claim 67, wherein the signal peptide comprises the amino acid of SEQ ID NO: 24, and wherein the amino acid is encoded by the nucleic acid sequence of SEQ ID NO: 25, or a nucleic acid sequence at least about 90% identical thereto.

69. The DNA vector of any one of claims 66-68, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 28, wherein the amino acid is encoded by a nucleic acid sequence of SEQ ID NO: 29, or a nucleic acid sequence at least 90% identical thereto.

70. The DNA vector of any one of claims 66-68, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises an E108M substitution (SEQ ID NO: 28).

71. The DNA vector of any one of claims 66-68, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises an E108M substitution, an N213Q substitution, and an N286Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 88).

72. The DNA vector of any one of claims 66-68, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises at least one amino acid substitutions selected from the group consisting of E108A, E108S, E108T, E108Q, E108M, E108K, E108L, E108R, E108N, E108D, E108G, E108H, E108I, E108F, E108P, E108W, E108Y, E108V, N213Q, N286Q, M384A, M384R, M384N, M384D, M384Q, M384E, M384G, M384H, M384I, M384L, M384K, M384F, M384S, M384T, M384Y, M384 V, L385V, L385K, L385A, L385N, L385H, L385S, L385T, N213Q, N286Q, and N413Q, relative to SEQ ID NO: 26 (SEQ ID NO: 85).

73. The DNA vector of any one of claims 30-32, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acidsequence further comprises an E108X substitution, wherein X = any amino acid relative to SEQ ID NO: 26 (SEQ ID NO: 86).

74. The DNA vector of any one of claims 30-32, wherein [B] comprises TNAPLP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises a N213Q substitution and a N286Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 87).

75. The DNA vector of any one of claims 30-32, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises an E108M substitution, an N213Q substitution, and an N286Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 88).

76. The DNA vector of any one of claims 30-32, wherein [B] comprises TNAPLP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises an E108A substitution relative to SEQ ID NO: 26 (SEQ ID NO: 89).

77. The DNA vector of any one of claims 30-32, wherein [B] comprises TNAPLP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises an E108A substitution, a N213Q substitution, and a N286Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 90).

78. The DNA vector of any one of claims 30-32, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises an E108X substitution, a N213X substitution, and a N286X substitution, wherein X = any amino acid relative to SEQ ID NO: 26 (SEQ ID NO: 91).

79. The DNA vector of any one of claims 28-30, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises a N213Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 95).

80. The DNA vector of any one of claims 30-32, wherein [B] comprises TNALP comprising an amino acid sequence of SEQ ID NO: 26, wherein the amino acid sequence further comprises an E108X substitution, a N286Q substitution relative to SEQ ID NO: 26 (SEQ ID NO: 96).

81. The DNA vector of any one of claims 66-80, wherein the Fc domain comprises an amino acid sequence of SEQ ID NO: 30, wherein the amino acid sequence is encoded by the nucleic acid sequence of SEQ ID NO: 31, or a nucleic acid sequence at least about 90% identical thereto.

82. The DNA vector of any one of claims 66-81, wherein [R] comprises an amino acid sequence of SEQ ID NO: 92, or an amino acid sequence at least about 90% identical thereto.

83. The DNA vector of any one of claims 66-81, wherein [R] comprises an amino acid sequence of SEQ ID NO: 93, or an amino acid sequence at least about 90% identical thereto.

84. The DNA vector of any one of claims 66-83, wherein [E] is the amino acid sequence DSS.

85. The DNA vector of claim 84, wherein y = 6, wherein [E]y comprises an amino acid sequence of SEQ ID NO: 32, wherein the amino acid sequence is encoded by the nucleic acid sequence of SEQ ID NO: 59, or a nucleic acid sequence at least about 90% identical thereto.

86. The DNA vector of any one of claims 66-83, wherein [E] is selected from aspartate, glutamate, glutamate-glutamate-serine (EES), or valine-histidine-histidine (VHH).

87. The DNA vector of any one of claims 66-86, wherein the promoter is an EF1L promoter.

88. The DNA vector of any one of claims 66-86, wherein the promoter comprises a nucleic acid sequence of SEQ ID NO: 33, or an amino acid sequence at least about 90% identical thereto.

89. The DNA vector of any one of claims 66-86, wherein the promoter comprises a nucleic acid sequence of SEQ ID NO: 34, or an amino acid sequence at least about 90% identical thereto.

90. The DNA vector of any one of claims 66-87, wherein the first portion further comprises an enhancer nucleic acid sequence.

91. The DNA vector of claim 81, wherein the enhancer nucleic acid sequence comprises SEQ ID NO: 35, or a nucleic acid sequence at least about 90% identical thereto.

92. The DNA vector of any one of claims 66-91, wherein the first portion further comprises a post-transcriptional regulatory element.

93. The DNA vector of claim 92, wherein the post-transcriptional regulatory element is a woodchuck post-transcriptional regulatory element (WPRE).

94. The DNA vector of claim 93, wherein the WPRE is selected from the group consisting of SEQ ID NO: 61, SEQ ID NO: 62, and a nucleic acid sequence at least about 90% identical thereto.

95. The DNA vector of any one of claims 66-94, wherein the first portion further comprises a polyadenylation (poly(A)) sequence.

96. The DNA vector of claim 95, wherein the poly(A) sequence is a bovine growth hormone poly(A) sequence selected from the group consisting of SEQ ID NO: 63, SEQ ID NO: 64, and a nucleic acid sequence at least about 90% identical thereto.

97. The DNA vector of any one of claims 66-96, wherein the first AAV-ITR is selected from the group consisting of SEQ ID NOS: 36, 38, 40, 42, 44, 46, 48, 50, 52, and a sequence at least about 90% identical thereto.

98. The DNA vector of any one of claims 66-97, wherein the first AAV-ITR is selected from the group consisting of SEQ ID NOS: 37, 39, 41, 43, 45, 47, 49, 51, 53, and a sequence at least about 90% identical thereto.

99. The DNA vector of any one of claims 66-98, wherein the Ori is selected from the group consisting of SEQ ID NOS: 54, 55, 56, 57, 58, and a sequence at least about 90% identical thereto.

100. The DNA vector of any one of claims 66-99, wherein the second portion comprises a nucleic acid sequence of SEQ ID NO: 13, or a sequence at least 90% identical thereto.

101. The DNA vector of any one of claims 66-99, wherein the second portion comprises a nucleic acid sequence of SEQ ID NO: 14, or a sequence at least 90% identical thereto.

102. The DNA vector of any one of claims 66-99, wherein the second portion comprises a nucleic acid sequence of SEQ ID NO: 15, or a sequence at least 90% identical thereto.

103. The DNA vector of any one of claims 66-99, wherein the second portion comprises a nucleic acid sequence of SEQ ID NO: 16, or a sequence at least 90% identical thereto.

104. The DNA vector of any one of claims 1-11, 16-61, and 66-99, wherein the Ori further comprises one or more additional DNA sequences selected from small runs ofextraneous and or spacer nucleotide sequences of from about 1 bp to about 20 bps, cloning or recombination sites, or a LoxP site, FRT site, attB and attP site, attL or attR sire, or alternative recombination target sites derived from Lox511 or Lox66 sites.

105. The DNA vector of any one of claims 1-104, wherein the DNA vector has less than about 4% of CpG motifs methylated.

106. The DNA vector of any one of claims 1-105, wherein the second portion comprising a nucleic acid sequence is capable of forming at least one specialized secondary structure.

107. The DNA vector of claim 106, wherein the first AAV-ITR sequence and the second AAV-ITR sequence are capable of aligning to form two double-stranded arms of at least about 80 basepairs (bps).

108. The DNA vector of claim 107, wherein the two double-stranded arms of at least 80 bps further comprise a loop formed at the end of each extended arm by the Ori.

109. A pharmaceutical composition suitable for administration to a human comprising: (i) a circular, non-viral, non-integrating DNA vector of any one of claims 1-108; and (ii) a pharmaceutically acceptable carrier or excipient.

110. The pharmaceutical composition of claim 109, wherein the pharmaceutical composition is formulated with a lipid-based delivery vehicle.

111. The pharmaceutical composition of claim 109, wherein the pharmaceutical composition is formulated as lipid nanoparticles.

112. The pharmaceutical composition of claim 109, wherein the pharmaceutical composition is formulated as lipid nanocapsules.

113. The pharmaceutical composition of claim 109, wherein the pharmaceutical composition is formulated with one or more polymers.

114. A method of treating a subject having hypophosphatasia (HPP) comprising administering to the subject a therapeutically effective amount of a DNA vector of any one of claims 1-108.

115. A method of treating a subject having hypophosphatasia (HPP) comprising administering to the subject a therapeutically effective amount of a pharmaceutical composition of any one of claims 109-114.

116. The method of claim 115, wherein the pharmaceutical composition is administered weekly, monthly, once every two months, once every six months, or yearly.

117. A pharmaceutical composition suitable for administration to a human for the treatment of HPP comprising: (i) a circular, non-viral, non-integrating DNA vector comprising a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 17-23, SEQ ID NOS: 97-98, and a nucleic acid sequence at least about 90% identical thereto; and (ii) a lipid nanoparticle.

118. The pharmaceutical composition of claim 117, comprising the nucleic acid sequence of SEQ ID NO: 17, or a nucleic acid sequence at least about 90% identical thereto.

119. The pharmaceutical composition of claim 118, comprising the nucleic acid sequence of SEQ ID NO:

17.

120. The pharmaceutical composition of claim 117, comprising the nucleic acid sequence of SEQ ID NO: 18, or a nucleic acid sequence at least about 90% identical thereto.

121. The pharmaceutical composition of claim 120, comprising the nucleic acid sequence of SEQ ID NO:

18.

122. The pharmaceutical composition of claim 117, comprising the nucleic acid sequence of SEQ ID NO: 19, or a nucleic acid sequence at least about 90% identical thereto.

123. The pharmaceutical composition of claim 122, comprising the nucleic acid sequence of SEQ ID NO:

19.

124. The pharmaceutical composition of claim 117, comprising the nucleic acid sequence of SEQ ID NO: 20, or a nucleic acid sequence at least about 90% identical thereto 125. The pharmaceutical composition of claim 124, comprising the nucleic acid sequence of SEQ ID NO: 20.

126. The pharmaceutical composition of claim 117, comprising the nucleic acid sequence of SEQ ID NO: 21, or a nucleic acid sequence at least about 90% identical thereto.

127. The pharmaceutical composition of claim 126, comprising the nucleic acid sequence of SEQ ID NO:

21.

128. The pharmaceutical composition of claim 117, comprising the nucleic acid sequence of SEQ ID NO: 22, or a nucleic acid sequence at least about 90% identical thereto.

129. The pharmaceutical composition of claim 128, comprising the nucleic acid sequence of SEQ ID NO:

22.

130. The pharmaceutical composition of claim 117, comprising the nucleic acid sequence of SEQ ID NO: 23, or a nucleic acid sequence at least about 90% identical thereto.

131. The pharmaceutical composition of claim 130, comprising the nucleic acid sequence of SEQ ID NO:

23.

132. The pharmaceutical composition of claim 117, comprising the nucleic acid sequence of SEQ ID NO: 97, or a nucleic acid sequence at least about 90% identical thereto.

133. The pharmaceutical composition of claim 117, comprising the nucleic acid sequence of SEQ ID NO:

97.

134. The pharmaceutical composition of claim 117, comprising the nucleic acid sequence of SEQ ID NO: 98, or a nucleic acid sequence at least about 90% identical thereto.

135. The pharmaceutical composition of claim 117, comprising the nucleic acid sequence of SEQ ID NO:

98.

136. A method of preventing or treating hypophosphatasia, comprising administering a therapeutically effective amount of the DNA vector of any one of claims 117-135, to a patient in need of treatment thereof.

137. Use of a pharmaceutical composition of any one of claims 117-135 in the treatment of hypophosphatasia.

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