Identity of zinc finger nucleases with specificity to herpes simplex virus type II genomic DNA: novel HSV-2 vaccine/therapy precursors

“In-vivo, as noted above, the zinc finger nucleases function as dimers (or pairs) [54-57].”

“[38]), constructs of 684 ZFNs (hybrid, chimeric Zif-FN) with specificity to the HSV-2 genome (see additional file 3) was attained.”

“Based on the fact that infection with HSV-2 is associated with an increased risk of HIV-1 acquisition and transmission [ 9 ,10], that existing treatment options for HSV-2 only serve to resolve the symptomatology (ulcers) associated with active-HSV-2 infection and do not eliminate latent-virus [13-15], and that there a…”

“In-vivo, as noted above, the zinc finger nucleases function as dimers (or pairs) [54-57].”

“In-vivo, as noted above, the zinc finger nucleases function as dimers (or pairs) [54-57].”

“The majority of recombinant-HSV-2-based vectors have yielded encouraging results in Phase I and Phase II clinical trials [21-24,47,48]; while genetically modified-HSV-I-based vectors raise safety-concerns around their potential to induce apoptosis in CNS neurons, causing severe and often fatal encephalitis or epilepti…”

“Therefore, treatment for HSV-2 and other genital ulcerative diseases [9] is an acceptable measure when considering reducing the risk of HIV-1 sexual transmission and acquisition [10-12].”

“This limits their clinical relevance to only treating ulcerative lesions of HSV-2 [ 14 ,15], rather than clearing the infection.”

“Alternatively, although several clinical trials concerning HSV based viral vectors have been conducted and demonstrated their safety [40-46], no clinical trial-data exists concerning the safety of our proposed and modeled HSV-2 viral vectors that deliver and transduce the diploid copy or pair of the HSV-2 genome-speci…”

“In-vivo, dimerization of ZFNs is mediated by the FokI cleavage domain through cleavage of a five or six base pair ‘spacer’ sequence that separates the two inverted target ‘half sites’ [34-36].”

“In-vivo, as noted above, the zinc finger nucleases function as dimers (or pairs) [54-57].”

“These are bracketed by inverted repeat sequences (TRL-IRL and IRS-TRS) that readily allow isomerization and recombination of the two regions [27,28].”

“In-vivo, dimerization of ZFNs is mediated by the FokI cleavage domain through cleavage of a five or six base pair ‘spacer’ sequence that separates the two inverted target ‘half sites’ [34-36].”

“Active infection with HSV-1 or HSV-2 leads to ulcerative lesions of the oral or genital mucosa, respectively [3,4]; latent infection with these viral species is largely asymptomatic.”

“Furthermore, a model concerning a recombinant (ICP10-PK mutant) replication competent HSV-2 viral vector for delivering and transducing a diploid copy (or pair) of the HSV-2-genome-specific ZFN genotype within neuronal tissue, is presented.”

“The majority of recombinant-HSV-2-based vectors have yielded encouraging results in Phase I and Phase II clinical trials [21-24,47,48]; while genetically modified-HSV-I-based vectors raise safety-concerns around their potential to induce apoptosis in CNS neurons, causing severe and often fatal encephalitis or epilepti…”

“The majority of the viral envelope glycoproteins (gD) are antigenically related to those of HSV-1, whereas gG1 and gG2 are type-specific [27, 29 ].”

“• Firstly, it was observed in accordance with Dudek and Knipe [42], that it is possible to use two types of replication-impaired HSV mutant viral strains as vectors for HSV vaccines, these being (a) replication-defective mutant strains and (b) single-cycle mutant strains.”

“[61], who have previously identified HSV-1 specific homing (mega-) endonucleases and demonstrated their inhibition of HSV-1 infection in cultured cells, shows that artificial mega-nucleases (zinc finger nucleases) mediated mutagenesis offers a novel therapeutic strategy for targeting HSV1 and episomal Hepatitis B Viru…”

“In-vivo, dimerization of ZFNs is mediated by the FokI cleavage domain through cleavage of a five or six base pair ‘spacer’ sequence that separates the two inverted target ‘half sites’ [34-36].”

“This array of numerous gene-products, many of which are indispensable for virus growth in vitro, underlies the efficacious virulence evolved by HSV-2 towards the evasion of host defenses including preventing apoptosis in the infected host cell, blockade of pathways for interferon-induction and production, and down-reg…”

“Existing biomedical interventions for HSV-2 treatment are only applicable to actively replicating HSV-2 particles [13].”

“sites for ZFNs were identified that are potentially best-suited for the complete excision of over 60% of HSV-2 genomic material from within infected latently human cells (see additional files 4 and 5, respectively, and Table 1) through the process of non-homologous end joining (NHEJ), following the introduction of a d…”

“The majority of recombinant-HSV-2-based vectors have yielded encouraging results in Phase I and Phase II clinical trials [21-24,47,48]; while genetically modified-HSV-I-based vectors raise safety-concerns around their potential to induce apoptosis in CNS neurons, causing severe and often fatal encephalitis or epilepti…”

“Furthermore, a model concerning a recombinant (ICP10-PK mutant) replication competent HSV-2 viral vector for delivering and transducing a diploid copy (or pair) of the HSV-2-genome-specific ZFN genotype within neuronal tissue, is presented.”

“Among the latter group of HSV-2 mutant vectors, the HSV-2 mutant ΔRR is deleted in the ICP10 RR domain for the enzyme ribonucleotide reductase, which is required for virus replication in neurons; the HSV-2 mutant ΔPK is deleted in only the protein kinase (PK) domain of the large subunit of ribonucleotide reductase (R1…”

“specified nucleotide sequence, zinc fingers have previously been used to direct small molecules into unique sequences within the human genome including genes of integrated viruses [58,59].”

“[37] concerning homing (mega-) endonucleases specific to HSV-1, ZF-nuclease-based disruption or abolition of HSV-2 genomes has yet to be reported.”

“This picture of a lifelong latent infection that can be reactivated presents unique challenges for the biomedical management of HSV-2 infections [18].”

“specified nucleotide sequence, zinc fingers have previously been used to direct small molecules into unique sequences within the human genome including genes of integrated viruses [58,59].”

“The majority of recombinant-HSV-2-based vectors have yielded encouraging results in Phase I and Phase II clinical trials [ 21-24 ,47,48]; while genetically modified-HSV-I-based vectors raise safety-concerns around their potential to induce apoptosis in CNS neurons, causing severe and often fatal encephalitis or epilep…”

“Therefore, treatment for HSV-2 and other genital ulcerative diseases [9] is an acceptable measure when considering reducing the risk of HIV-1 sexual transmission and acquisition [10-12].”

“Alternatively, although several clinical trials concerning HSV based viral vectors have been conducted and demonstrated their safety [40-46], no clinical trial-data exists concerning the safety of our proposed and modeled HSV-2 viral vectors that deliver and transduce the diploid copy or pair of the HSV-2 genome-speci…”

“Alternatively, although several clinical trials concerning HSV based viral vectors have been conducted and demonstrated their safety [40-46], no clinical trial-data exists concerning the safety of our proposed and modeled HSV-2 viral vectors that deliver and transduce the diploid copy or pair of the HSV-2 genome-speci…”

“The final construct, denoted as HSV-2ΔRR [ICP10PK- 2xZifHSV-2-FN] would contain ICP10PK- 2xZifHSV-2-FN driven by the authentic ICP10 promoter, which is regulated by IE kinetics (independent of virus replication) and responds to AP-1 [25, 52 ,53].”

“There have been substantive efforts to develop an efficacious HSV-2 vaccine based on existing biomedical evidence, and these are on-going [19,20].”

“[60] demonstrated that several HSV-2 mutants with cumulative restriction site deletions in one of the repeats (TRL or IRL) have a measurable growth disadvantage relative to wild-type virus in tissue culture.”

“Based on the fact that infection with HSV-2 is associated with an increased risk of HIV-1 acquisition and transmission [9,10], that existing treatment options for HSV-2 only serve to resolve the symptomatology (ulcers) associated with active-HSV-2 infection and do not eliminate latent-virus [13-15], and that there are…”

“Alternatively, although several clinical trials concerning HSV based viral vectors have been conducted and demonstrated their safety [40-46], no clinical trial-data exists concerning the safety of our proposed and modeled HSV-2 viral vectors that deliver and transduce the diploid copy or pair of the HSV-2 genome-speci…”

“Alternatively, although several clinical trials concerning HSV based viral vectors have been conducted and demonstrated their safety [40-46], no clinical trial-data exists concerning the safety of our proposed and modeled HSV-2 viral vectors that deliver and transduce the diploid copy or pair of the HSV-2 genome-speci…”

“There have been substantive efforts to develop an efficacious HSV-2 vaccine based on existing biomedical evidence, and these are on-going [19,20].”

“genital ulcerations following reactivation during or following a bout of immune suppression caused by a common cold, after infection with HIV or chemotherapy treatment, among others [16,17].”

“Based on the fact that infection with HSV-2 is associated with an increased risk of HIV-1 acquisition and transmission [9,10], that existing treatment options for HSV-2 only serve to resolve the symptomatology (ulcers) associated with active-HSV-2 infection and do not eliminate latent-virus [13-15], and that there are…”

“Modeling the prototype recombinant HSV-2 viral vector delivering and transducing HSV- 2 genome-specific-ZFN genotype A review of available relevant literature [25, 41-50 ] concerning HSV-2 viral vectors was carried out, as is sequentially described in the respective results’ sub-section above.”

“genital ulcerations following reactivation during or following a bout of immune suppression caused by a common cold, after infection with HIV or chemotherapy treatment, among others [16,17].”

“Delivery of the HE to the neurons where HSV is latent could be achieved by using replication-incompetent HSV-based delivery vectors (200), which have been developed for gene therapy of diseases of the nervous system (62).”

“ZFN technology has also been used to successfully target proviral DNA in HBV [49], HSV-2 [50] and HTLV-1 [51] infection.”

“Unknown Bullous pemphigoid Characterized by large, tense, subepidermal bullae, involving the groin, axillae, trunk, thighs, and forearms [139] The decreased serum level of Zn has been reported [147] Unknown Epidermolysis bullosa A group of rare genetic disorders, common in the formation of blisters in response to mino…”

“[31,32], our model HPV-binding ZFNs would be usable first to cleave and disrupt HPV type 16 genomic DNA at the contextual positions corresponding to about 0.”