Category Archives: Gene therapy

NGN-401, Neurogenes gene therapy candidate for Rett syndrome, was well tolerated in the first three patients dosed in a Phase 1/2 clinical trial.

The Phase 1/2 trial (NCT05898620) is enrolling girls with Rett, ages 4-10, at three sites in the U.S. The company also plans to establish the first clinical trial for the treatment in the U.K., following recent regulatory approval.

Trial results were presented during the American Society of Gene and Cell Therapy (ASGCT) annual meeting, held May 7-11 in Baltimore. The company expects to share interim efficacy data from the first group of patients later this year, and additional results, including data from a second group of participants, in 2025.

The NGN-401 datapresented at ASGCT demonstrate a favorable tolerability profile in the first three pediatric patients with no signs or symptoms of overexpression-related toxicity reported in any patient, Rachel McMinn, PhD, Neurogenes founder and CEO, said in a company press release.

In most cases, Rett syndrome is caused by mutations in the MECP2 gene that impair the function of MeCP2, a protein that regulates the activity of other genes. The loss of MeCP2 impairs the growth and connectivity of neurons (nerve cells), leading to disease symptoms.

NGN-401 provides a full-length version of the gene, packaged in a harmless adeno-associated virus (AAV). The therapy is delivered directly into the fluid-filled cavities of the brain, a process known as intracerebroventricular infusion, in a one-time treatment.

The gene therapy was developed using Neurogenes EXACT gene regulation technology, for the delivery of highly controlled and consistent levels of the lab-made MECP2 gene without the toxic effects associated with high levels of the MeCP2 protein, or overexpression, associated with conventional gene therapy.

We designed NGN-401 to overcome the limitations of conventional gene therapy for Rett syndrome by incorporating our EXACT transgene regulation technology, which we believe provides tolerable and therapeutic levels of protein expression to the key areas of the brain and nervous system that drive disease, McMinn said.

Preclinical studies showed NGN-401 extended survival and eased the disease burden in a mouse model of Rett syndrome, without signs of toxicity associated with MeCP2. Data presented at ASGCT showed that the therapy was well tolerated in non-human primates, without overexpression concerns.

The ongoing clinical trial is assessing NGN-401s safety, tolerability, and preliminary efficacy in girls with Rett syndrome, following them for several years after treatment. The first group of patients consists of eight girls, receiving a low NGN-401 dose (1x1E 15 vector genomes).

The first two girls were dosed in 2023, and the third received the treatment this year. So far, the therapy has been well tolerated, with no treatment- or procedure-related serious adverse events, after a follow-up between three and nine months.

All reported adverse events were mild, and were mainly known potential risks of AAV, including increased blood levels of liver enzymes and decreased levels of certain immune proteins.

There were no signs of overexpression toxicity, even in a girl with a milder form of Rett, predicted to result in residual MECP2 gene activity.

A second group of eight girls will receive a high dose of NGN-401 (3 x 10^15 vector genomes). As with the first group, the first three patients will be dosed in a staggered manner, and if the gene therapy is considered safe, the other five will be dosed in parallel.

Efficacy measures will include clinician-rated assessments of symptom severity and patient improvement, as well the Rett Syndrome Behavior Questionnaire that evaluates behavioral challenges.

Gene therapy has the potential to address the underlying cause of Rett syndrome with a one-time treatment, and these interim safety data from the NGN-401 trial provide an important milestone on the path to realizing its potential for patients, said Bernhard Suter, MD, medical director of the Blue Bird Circle Rett Center at Texas Childrens Hospital and principal investigator in the clinical trial.

Read more:
Rett gene therapy NGN-401 safe in first patients dosed in trial - Rett Syndrome News

A CRISPR-based gene-editing therapy called EBT-101 was safe and well tolerated but did not prevent viral rebound in three participants who stopped antiretroviral treatment in an early study, according to a presentation last week at the American Society of Gene & Cell Therapy annual meeting.

As aidsmap previously reported, researchers presented findings at a conference last October showing that EBT-101 was well distributed in the body and did not cause serious side effects in the first three treated study participants. Although the presentation did not include data about whether the treatment actually worked to control HIV, that didnt stop the Daily Mail from proclaiming that a cure for HIV could be months away one of the many exaggerated claims over the years about the state of HIV cure research.

A unit of heredity, that determines a specific feature of the shape of a living organism. This genetic element is a sequence of DNA (or RNA, for viruses), located in a very specific place (locus) of a chromosome.

To eliminate a disease or a condition in an individual, or to fully restore health. A cure for HIV infection is one of the ultimate long-term goals of research today. It refers to a strategy or strategies that would eliminate HIV from a persons body, or permanently control the virus and render it unable to cause disease. A sterilising cure would completely eliminate the virus. A functional cure would suppress HIV viral load, keeping it below the level of detection without the use of ART. The virus would not be eliminated from the body but would be effectively controlled and prevented from causing any illness.

A type of experimental treatment in which foreign genetic material (DNA or RNA) is inserted into a person's cells to prevent or fight disease.

But those data are out now, and the news is generally unfavourable. EBT-101 did not maintain HIV viral suppression when used alone at the initial dose tested, though it may have delayed viral rebound in one participant. Also, its good safety profile suggests that similar CRISPR approaches may be feasible for other latent viral infections such as herpes simplex and hepatitis B.

We know that many people were hopeful that a first trial could provide evidence of a possible cure for HIV because the field has been waiting over 20 years for a cure, Excision BioTherapeutics senior vice president Dr William Kennedy said in a news release. However, it was essential that this clinical trial establish safety for EBT-101 as a gene therapy product as well as safety related to the use of CRISPR for the field.

Antiretroviral therapy can keep HIV replication suppressed indefinitely, but the virus inserts its genetic blueprints into the DNA of human cells and establishes a long-lasting reservoir that the drugs cant reach. This integrated HIV DNA lies dormant in resting T cells during treatment, but it can start producing new virus when antiretrovirals are stopped, making a cure nearly impossible. The only way to determine whether an experimental intervention leads to long-term remission is to discontinue antiretroviral therapy with careful monitoring in an analytic treatment interruption.

Professor Kamel Khalili of Temple University in Philadelphia and colleagues have been studying gene therapy with the aim of curing HIV for more than a decade. Their work employs CRISPR/Cas9 sometimes referred to as molecular scissors a technology that combines guide RNAs that home in on specific segments of DNA and a nuclease enzyme that cuts the genetic material at the desired site.

In 2014 and 2016, the researchers reported that a CRISPR/Cas9 tool could cut out HIV genes from CD4 cells in laboratory studies. A study published in 2019 showed that this approach could remove integrated HIV genes and clear latent viral reservoirs in mice. And at the 2019 Conference on Retroviruses and Opportunistic Infections, the Temple University team reported that CRISPR/Cas9 therapy successfully removed segments of an HIV-like virus from viral reservoirs in monkeys.

This research led to the development of EBT-101, a CRISPR-based therapy delivered by an adeno-associated virus vector that uses two guide RNAs to target three sites on the integrated HIV genome. Making cuts at these locations prevents the production of intact new virus. Last August, researchers reported that a single dose of a simian version of the therapy safely and effectively removed integrated SIV in monkeys on antiretroviral therapy. But this study did not include a treatment interruption, so it could not show whether the animals were functionally cured.

The first human clinical trial of EBT-101 (NCT05144386) started in 2022, testing the therapy in people on antiretroviral treatment with a stable undetectable viral load. Excision announced that the first participant in the phase I/II trial received EBT-101 that July, and the study protocol called for participants who maintained viral suppression at 12 weeks after receiving the gene therapy to undergo an analytic treatment interruption.

At last weeks conference, Dr Rachel Presti of Washington University St. Louis School of Medicine provided updated study results. Of the five participants who received a single infusion of the initial dose of EBT-101, three stopped antiretroviral therapy. All three experienced viral rebound and had to restart antiretrovirals. This likely occurred because the gene therapy did not reach all cells harbouring latent HIV, and even a very small number of cells containing residual HIV DNA is enough to re-establish viral replication.

One EBT-101 recipient was able to maintain viral suppression for 16 weeks after treatment discontinuation, considerably longer than it typically takes for the virus to rebound after stopping antiretrovirals. This suggests that EBT-101 or similar CRISPR therapies might one day play a role in a combination cure strategy.

Initial data from the EBT-101-001 trial provides important clinical evidence thatagene editingtreatment modality can be safely delivered fortargetingthe HIV DNA reservoirs in human cells, Presti said. This study provides researchers with invaluable insights for how CRISPR technology can be applied for addressing infectious disease and was an important first step towards additional programs designed to optimize this treatment modality for treating the millions of individuals who are impacted by HIV and other infectious disease.

Excision is now testing a higher dose of EBT-101 in a second cohort and is exploring new CRISPR delivery methods that might be more efficient than the adeno-associated virus vector. One possibility is lipid nanoparticles like the ones used to deliver messenger RNA in COVID-19 vaccines.

Viral rebound likely occurred because the gene therapy did not reach all cells harbouring latent HIV

The company is also exploring CRISPR-based approaches for other latent infections. In other presentations at last weeks meeting, researchers reported promising preclinical results for experimental therapies for herpes simplex (EBT-104) and hepatitis B (EBT-107). Herpes simplex virus (HSV) persists in nerve cells, from which it can reactivate to cause cold sores, genital herpes and keratitis (eye inflammation). Hepatitis B virus (HBV) establishes chronic infection in the liver, where it can lead to cirrhosis and liver cancer. Unlike HIV and other retroviruses, however, HSV and HBV do not integrate their genetic blueprints into the chromosomes of host cells, so they may be easier to remove.

Many lessons have been learned from the small number of people who naturally control HIV, the somewhat larger group of post-treatment controllers and the handful of people who have been cured after stem cell transplants. But for now, a broadly applicable functional cure remains a long-term prospect.

See more here:
CRISPR gene therapy EBT-101 does not prevent HIV viral rebound - aidsmap

WATCH TIME: 5 minutes

"Another issue with gene therapy is choosing the proper patient that might be best for a particular gene therapeutic. Then [figuring out] how do you deliver that gene therapy to the brain, most of the patients to date have had that done surgically."

In the current realm of treatments for Parkinson disease (PD), clinicians and patients living with the disease highlight the need for more innovative approaches to address the progressive decline in dopamine network function from the condition. As a promising avenue, gene therapy has emerged to offer the potential for sustained restoration of neuronal function by introducing genetic material to regulate dopamine levels and improve dopaminergic signaling.1 Thus, the clinical gene therapy trials in progress for PD are primarily aimed towards enhancing dopamine production enzymatically and reinstating the integrity of the nigrostriatal pathway.

Despite the potential of gene therapy for PD, substantial challenges impede its translation into clinical practice. The blood-brain barrier presents an obstacle for this since it impedes gene delivery into the central nervous system (CNS) unless it is directly administered. Modified vectors, both viral and non-viral, are in development to enhance delivery efficiency to target these locations in the CNS; however, the limited approved gene therapy approaches for CNS diseases like PD reflect the ongoing safety concerns associated with these viral vectors.2 To combat this, there has been an increasing focus on developing alternative non-viral biomaterial vectors to mitigate these safety risks in CNS disorders such as PD.

Michael Kaplitt, MD, PhD, professor of neurological surgery and vice chairman for research in the department of neurological surgery at Weill Cornell Medicine, recently sat down with NeurologyLive in an interview to discuss the regulatory barriers that exist for the approval of gene therapies, and how recent approvals in other neurological disorders have influenced the pathway for PD treatments. He also talked about the challenges associated with conducting long-term safety studies for gene therapies compared with conventional drugs. In addition, Kaplitt spoke about how patient selection and delivery methods can impact the effectiveness and feasibility of gene therapy for PD.

View post:
Regulatory Pathways and Clinical Trials for Gene Therapy in Parkinson Disease: Michael Kaplitt, MD, PhD - Neurology Live

Meganuclease therapy reduces ganglionic viral load after ocular or genital HSV infection

We previously evaluated several AAV serotypes for delivery of meganucleases to latently infected mice, and found the best results with AAV-Rh10, followed by AAV8 and AAV18. To further improve efficacy, we tested additional neurotropic AAV serotypes, including AAV7, AAV9, AAV-DJ, and AAV-DJ/810,11, for delivery of the anti-HSV1 meganuclease, m5, at a dose of 1012 AAV genomes (vg) per mouse (Fig.S1a), using our model of orofacial HSV disease. Both AAV9 and AAV-Dj/8 were superior to 1012 vg AAV-Rh10, the best of our previously used serotypes8, showing HSV reductions in superior cervical ganglia (SCG) of 95% (p=105) and 90% (p=0.018), respectively, relative to untreated controls (Fig.S1b), comparing favorably with the 65% reduction previously obtained with m5 alone delivered with AAV-Rh10 8. Similarly, AAV9 and AAV-Dj/8 showed better activity than AAV-Rh10 in trigeminal ganglia (TG), with HSV load reductions of 48% (p=0.07) and 41% (p=0.5), respectively (Fig.S1b), compared with our prior observation of no detectable reduction using AAV-Rh10 delivering m58. The route of AAV administration (retro-orbital vein vs. intradermally into the whisker pad) did not have any detectable impact on either AAV transduction or gene editing efficiencies (Fig.S1eg).

We previously demonstrated that gene editing of HSV could be increased by using combinations of AAV serotypes for meganuclease delivery, rather than a single AAV serotype, a finding we ascribed to the heterogeneity of neuronal subsets within HSV-infected ganglia8. We therefore evaluated gene editing with the anti-HSV1 meganuclease m5, which cleaves a sequence in the UL19 gene coding for the major capsid protein VP512, when delivered using single AAV serotypes vs. combinations of AAV9, AAV-Dj/8, and AAV-Rh10 (Fig.S2a) administered as a total dose of 1012 vg per mouse. In agreement with our previous results, combinations of AAV serotypes led to robust HSV gene editing, with the triple combination of AAV9, AAV-Dj/8, and AAV-Rh10 showing especially strong reductions in HSV loads and mutagenesis of residual HSV across both SCG and TG (Fig.S2bg).

While orofacial infections with HSV are extremely common, genital infections, which lead to latent infection of dorsal root ganglia (DRG), also represent a major cause of morbidity. We therefore established latent genital infections in mice by intravaginal inoculation with HSV-1 after treatment with Depo Provera, which synchronizes the estrus cycle and increases HSV infection13. Infected mice were treated with a total dose of 31012 vg of the AAV9, AAV-Dj/8, and AAV-Rh10 combination delivering two HSV1-specific meganucleases simultaneously (m5 along with m8, which targets a sequence in the UL30 gene coding for the catalytic subunit of the viral DNA polymerase12. In parallel, we tested the same AAV combination against latent orofacial HSV infection as described above (Fig.1a, b). Remarkably, efficacy in the vaginal model of infection was the highest we have observed to date, with a 97.7% reduction in HSV viral load in DRG (Fig.1c). This compared favorably with the orofacial infection group treated in parallel, in which (in agreement with our previous studies) we observed robust gene editing with significant reductions of ganglionic HSV loads of 89% in SCG and 61% in TG (Fig.1d, e).

a, b Experimental timeline of (a) vaginal or (b) ocular infection and meganuclease therapy. RO, retroorbital; TV, tail vein. c HSV loads in DRGs from control (n=7) and dual meganuclease-treated (n=4) mice vaginally infected with HSV-1; p=0.001. d. HSV loads in SCGs and TGs from control (n=10) and dual meganuclease-treated (n=10) mice ocularly infected with HSV-1; p=0.0046 and 0.0034 for SCG and TG, respectively. e Gene editing at the m5 target site of residual virus quantified by T7E1 assay in SCG and TG from dual meganuclease-treated mice (n=10). Each graph shows individual and mean values with standard deviation, percent decrease of HSV loads in treated mice compared to control mice and statistical analysis (unpaired one-tailed Mann-Whitney test with **p<0.01). AAV loads are shown in Supplemental Fig.9a, b. Source data are provided as a Source Data file.

Mice generally show little if any spontaneous HSV reactivation, with minimal to no viral shedding at peripheral sites, limiting their utility in cure studies. The BET (Bromo and Extra-Terminal domain) bromodomain inhibitor JQ1 was reported to reactivate latent HSV in vitro in primary neuronal cultures, and HSV could be detected in the eyes of about one-third of latently infected mice treated with JQ114. To evaluate the utility of JQ1 for our cure work, we extended these studies to determine the quantitative kinetics of HSV shedding after JQ1 therapy.

A single intraperitoneal (IP) injection of JQ1 (50mg/kg) given to latently infected mice (Fig.S3a) led to detectable shedding from the eyes of 56% (5/9) of animals, compared with 0/9 animals treated with vehicle alone (Fig.S3b, c). Viral shedding was transient, peaking at 2 days post JQ1, with maximal viral loads ranging from about 102 to almost 106 copies/swab (Fig.S3c). A direct comparison suggested that JQ1 may be a more powerful reactivation stimulus for HSV than hyperthermic stress (HS)15, which in our hands led to detectable virus shedding in less than 20% of animals (2/12 HS vs 4/10 JQ1), with peak shedding viral loads two logs lower than after JQ1 treatment (Fig.S3df). Sequential treatment with JQ1 at one-week intervals led to repeated shedding episodes with similar kinetics as observed above (Fig.S4ae). Over the course of three sequential JQ1 reactivations (Fig.S4e), shedding from individual mice was stochastic; 10/12 (83%) mice shed detectable virus at least once, but only 1/12 (8%) shed after all three treatments, while 4/12 (33%) and 5/12 (42%) shed only after two or one of the three treatments, respectively (Fig.S4e and TableS1). Shedding was typically unilateral (only detected in one eye), despite the initial inoculation being to both eyes (unilateral shedding was observed in 33/37 (89%) of events). The side of shedding in one episode was not predictive of the side of future shedding events (TableS1). Importantly for cure studies, repeated weekly reactivation of virus with JQ1 up to 7 weekly injections did not change ganglionic viral loads compared with control animals (Fig.S4g, h).

The ability to reproducibly induce HSV reactivation and shedding with JQ1 allowed us to investigate the relationship between ganglionic viral load reduction using meganucleases and subsequent viral shedding at peripheral sites. Latently infected mice were treated as above using the AAV9, AAV-Dj/8, and AAV-Rh10 combination delivering HSV1-specific meganucleases m5 and m8 at a total dose of 31012 vg, or left untreated as controls. One month later, mice were administered JQ1, and eye swabs were collected daily for 4 days (Fig.2a). Consistent with our previous results, ganglionic tissues from treated mice showed a 98% and 42% reduction in mean viral loads in SCG and TG, respectively, when compared to control untreated animals (Fig.2b, c) and gene editing in the remaining viral genomes (Fig.2d). After JQ1 administration, only 3/10 (30%) of the dual meganuclease-treated mice had detectable virus in eye swabs, compared with 5/10 (50%) of control untreated animals (Fig.2e, f). The mean titer of HSV in positive eye swabs was 3104 copies/ml in the meganuclease-treated animals, compared with 1.2105 copies/ml in the control animals. Area under the curve analysis (AUC) demonstrated a 95% reduction (p=0.15) in total viral shedding in treated vs. control animals (Fig.2g). In a separate experiment performed similarly (Fig.3a), ganglionic tissues from treated mice showed 97% reduction in mean latent HSV genomes in both SCG and TG when compared to control untreated animals (Fig.3b, c) and gene editing in the remaining genomes (Fig.3d). While 3/8 mice from the control group shed virus with a mean viral titer of 8.2105 copies/ml, 0/8 meganuclease-treated mice had detectable shedding, representing a 100% decrease in total virus shed (p=0.10, Fig.3eg).

Experimental timeline of ocular infection, meganuclease treatment and viral reactivations with JQ1. a Experiment 1 (n=10 per group). HSV loads in SCGs (b: p=0.0057) and TGs (c). Percent decrease of HSV loads in treated mice compared to control mice and statistical analysis (unpaired one-tailed MannWhitney test with ns: not significant, **p<0.01) are indicated. Gene editing at the m5 target site of residual virus quantified by T7E1 assay in SCG and TG from dual meganuclease-treated mice (d). HSV titers in eye swabs collected daily from day 1 to 4 post JQ1 reactivation from control (e) and dual meganuclease-treated (f) infected mice. Panels 2i-k show data for both SCG and both TG from each mouse. Area under the curve (AUC) analysis (g) with p value (unpaired one-tailed MannWhitney test). AAV loads are shown in Supplemental Fig.9c, d. Each graph shows individual and mean values with standard deviation. Source data are provided as a Source Data file.

Experimental timeline of ocular infection, meganuclease treatment and viral reactivations with JQ1. a Experiment 2 (n=8 per group). HSV loads in SCGs (b) and TGs (c). Percent decrease of HSV loads in treated mice compared to control mice and statistical analysis (unpaired one-tailed Mann-Whitney test with ns: not significant, **p<0.01) are indicated. Gene editing at the m5 target site of residual virus quantified by T7E1 assay in SCG and TG from dual meganuclease-treated mice (d). HSV titers in eye swabs collected daily from day 1 to 4 post JQ1 reactivation from control (e) and dual meganuclease-treated (f) infected mice. Panels 2b-d show data for both SCG and both TG from each mouse. Area under the curve (AUC) analysis (g) with p value (unpaired one-tailed MannWhitney test). AAV loads are shown in Supplemental Fig.9e, f. Each graph shows individual and mean values with standard deviation. Source data are provided as a Source Data file.

AAV-vectored therapies are generally considered safe. Nevertheless, dose-limiting liver toxicity has been observed after AAV administration in humans, non-human primates, and mice, typically at doses of 21014 vg/kg or greater. The 31012 vg/animal dose (~11014 vg/kg) used in the experiments described in Figs.13 approached the level associated with liver toxicity in previous studies. Across multiple studies we observed that 7/70 (10%) animals treated with the 31012 vg/animal dose exhibited clinical signs consistent with hepatotoxicity, including weight change, bloating, and general health decline. Hepatotoxicity was confirmed in these animals by subsequent histopathological evaluation (Fig.S5 and TableS2). We therefore evaluated lower total doses of triple AAV serotype/dual meganuclease therapy (0.6, 1.2, or 1.81012 vg/animal or 1.8, 3.6, or 5.41013 vg/kg) for their tolerability and effects on viral load and JQ1-induced HSV shedding (Fig.4a). These doses showed substantially improved tolerability, both clinically and upon histopathological examination and quantification of the number of inflammatory cell foci (ICF) in livers (Fig.S6a). Dose-dependent reductions in ganglionic HSV loads were observed across the three treatment groups compared to controls, ranging from 69% and 47% in SCG and TG, respectively, at the 0.61012 dose to 94% and 73% at the 1.81012 dose (Fig.4b, c). To evaluate the effect of these reduced doses on HSV shedding, treated mice were subjected to three weekly rounds of JQ1 administration, followed by eye swabbing as described above. While the percentage of dual meganuclease-treated animals shedding virus after the first JQ1 reactivation was not reduced compared with the control mice, it was substantially lower than controls at all doses by the third JQ1 reactivation (0% (0/12), 8% (1/12) and 0% (0/12) for 0.6, 1.2, and 1.81012 vg/mouse groups, respectively, versus 18% (2/11) in the control group) (Fig.4fi). This finding that may relate to the two additional weeks available for meganuclease expression and gene editing activity by the time of the third JQ1 reactivation. Consistent with this interpretation, the reduction in total viral shedding, as determined by AUC analysis, appeared to become more complete over time, with up to a 97100% reduction in all three groups by the final JQ1 reactivation (Fig.4jl). The efficacy of reduced-dose dual meganuclease therapy (1.81012 vg) was confirmed in a separate experiment (Fig.5a), showing a significant decrease in ganglionic viral loads in both SCG and TG (Fig.5b, c). In this experiment, 7 of 12 control animals showed detectable viral shedding after JQ1 reactivation, compared with only 1 of 12 animals treated with AAV-meganuclease therapy, (Fig.5d, e) and reduction in total viral shedding, as determined by AUC analysis (Fig.5f, g). While none of the treated mice exhibited any clinical signs of hepatotoxicity, we did observe higher numbers of ICF in liver of treated animals receiving the 1.81012 vg dose compared to control mice (Fig.S6b). Histologic analysis of H&E stained TG sections from both control and treated animals revealed subtle evidence of neuronal injury, manifesting as neuronal degeneration, necrosis, and axonopathy. The scores grading prevalence and severity of the microscopic changes were higher in treated animals compared to control mice (Fig.S7 and TableS3). However, no mice in either the control or experimental group showed detectable signs of neuropathy.

a Experimental timeline of ocular infection, meganuclease treatment and viral reactivations with JQ1. b, c HSV loads and d, e, AAV loads in SCGs (b; p=0.0016, 0.0012, and <0.0001 for 0.6, 1.2, and 1.81012, respectively, d) and TGs (c; p=0.068, 0.0025 and 0.0016 for 0.6, 1.2, and 1.81012, respectively, e) in control infected mice (n=11) and infected mice treated with dual therapy delivered with 0.6 (n=12), 1.2 (n=12), or 1.8 (n=12)1012 total vg AAV dose. Percent decrease of HSV loads in treated mice compared to control mice and statistical analysis (ordinary one-way Anova, multiple comparisons with ns: not significant, **p<0.01, ****p<0.0001) are indicated. fi Virus titers in eye swabs collected daily from day 1 to 4 after each weekly JQ1 reactivation from control infected mice (f) and infected mice treated with dual therapy delivered with 0.6 (g), 1.2 (h), or 1.8 (i) 1012 total vg AAV dose. jl Area under the curve (AUC) analysis of virus shedding after first (j), second (k), and third (l) JQ1 reactivation from control infected mice (n=11) and infected mice treated with dual therapy delivered with 0.6 (n=12), 1.2 (n=12) or 1.8 (n=12)1012 total vg AAV dose. p values (unpaired, ordinary one-way Anova, with multiple comparisons) compared virus shedding between treatment groups and the control group. Each graph shows individual and mean values with standard deviation. Source data are provided as a Source Data file.

a Experimental timeline of ocular infection, meganuclease treatment and viral reactivations with JQ1. b HSV loads in SCGs (b; p=0.0012, and <0.0001 for CTRL vs AAV/MN no JQ1 and for CTRL vs AAV/MN 2x JQ1, respectively), and TGs (c; p=0.0089, and 0.0293 for CTRL vs AAV/MN no JQ1 and for CTRL vs AAV/MN 2x JQ1, respectively) of control infected mice either unreactivated (CTRL no JQ1) or reactivated (CTRL 2x JQ1) and infected mice treated with dual therapy delivered with 1.81012 total AAV dose either unreactivated (AAV/MN no JQ1) or reactivated (AAV/MN 2x JQ1), with n=12 per group. Percent decrease of HSV loads in treated mice compared to control mice and statistical analysis (unpaired one-tailed Mann-Whitney test with *p<0.05; **p<0.01, ****p<0.0001) are indicated. d, e Virus titers in eye swabs collected daily from day 1 to 4 after two weekly JQ1 reactivations (red arrows) from control infected mice (d) and infected mice treated with dual therapy delivered with 1.81012 total AAV dose (e). f, g Area under the curve (AUC) analysis after the first (f), and the second (g) JQ1 reactivation from control infected mice either unreactivated (CTRL no JQ1) or reactivated (CTRL 2x JQ1) and infected mice treated with dual therapy delivered with 1.81012 total AAV dose either unreactivated (AAV/MN no JQ1) or reactivated (AAV/MN 2x JQ1), with n=12 per group. p values (unpaired one-tailed MannWhitney test) are indicated. Each graph shows individual and mean values with standard deviation. AAV loads are shown in Supplemental Fig.9gh. Source data are provided as a Source Data file.

As noted above, genital HSV infection is a major cause of morbidity in humans. We therefore evaluated the reduced-dose dual meganuclease therapy (total dose of 1.81012 vg/animal) in vaginally infected mice (Fig.6a). In agreement with our previous results, the reduced-dose therapy led to a 78.8% (p=0.02) to 95.6% (p=0.006) reduction in latent virus genomes in DRGs (Fig.6b).

a Experimental timeline of intravaginal HSV-1 infection, meganuclease treatment and viral reactivations with JQ1. b HSV loads in DRGs from control infected mice reactivated with 3 weekly JQ1 injections and infected mice treated with dual therapy unreactivated, or reactivated with 3 weekly JQ1 injections with n=8 per group; p=0.0055, and 0.0198 (ordinary one-way Anova, multiple comparisons) for CTRL+JQ1 vs AAV/MN no JQ1 and for CTRL+JQ1 vs AAV/MN+JQ1, respectively. c, d HSV titers in vaginal swabs collected daily from day 1 to 4 post JQ1 injections (red arrows) from control (c) and dual meganuclease-treated (d) infected mice. Area under the curve (AUC) analysis after the first (e), second (f), and third (g) JQ1 reactivation from control infected mice (n=8) and infected mice treated with dual therapy, both reactivated with 3 weekly JQ1 injections (n=8). p values (unpaired one-tailed MannWhitney test) are indicated. Each graph shows individual and mean values with standard deviation. The AAV viral loads are shown in Supplemental Fig.9i. Source data are provided as a Source Data file.

We then sought to evaluate whether JQ1 could induce HSV shedding in the genital infection model, as we previously observed in the ocular infection model. Over 3 sequential JQ1 reactivations, only 2 of 8 control animals (and 1 of 8 AAV/meganuclease-treated animals) shed detectable virus, a rate lower than the 4050% reactivation we typically observe after ocular infection (Fig.6c, d). The apparently lower rate of reactivation seen in the vaginal model compared to the ocular model may be due to lower levels of ganglionic HSV loads in the DRG (102103 vg/106 cells in DRG, Fig.6b vs 104105 vg/106 cells in SCG or TG, Fig.5b, c). While this lower reactivation rate prohibited meaningful statistical analysis, the observation that 2 out of the 8 control mice shed virus over 2 to 3 sequential days, while only 1 of the 8 AAV-treated mice shed virus, on a single day and at a substantially lower level, is qualitatively in agreement with our observations after ocular infection (Fig.6cg).

The stochastic nature of clinical HSV reactivation16, recapitulated when induced by JQ1 in mice (Fig.S4 and TableS1), makes evaluation of viral shedding extremely resource-intensive. Practical constraints, including the number of animals that can be housed and studied at the same time, along with the extended duration of each study (~3 months), limited the statistical power of our individual experiments. We therefore performed a meta-analysis of data from all experiments presented above (Figs.16), combining evidence from infection sites (orofacial or genital), thus comparing 174 swabs from AAV/meganuclease-treated animals to 99 swabs from experimentally-matched controls. The primary endpoint was viral shedding, expressed either as a binary variable (equal to 1 for samples in which HSV was detected and 0 otherwise) or the log10-transformed AUC for quantitative viral shedding. The experiments depicted in Figs.16 represent all of the shedding studies with the dual meganuclease/triple AAV therapy we have performed as of this writing, and each suggests a strong and consistent trend toward a substantial reduction in viral shedding after AAV/meganuclease therapy. Across all studies, the proportion of swabs with detectable HSV was 48% lower among AAV/meganuclease-treated animals compared to controls. The meta-analysis confirmed that animals receiving AAV/meganuclease therapy had a statistically significant reduction in viral shedding (OR=0.41, p=0.010, by generalized linear mixed models, GLMM).

We then asked whether dose or duration of meganuclease therapy was associated with the probability of viral shedding (expressed as a binary variable) or the quantity of viral shedding (expressed as the log10-transformed AUC). Overall, the probability of viral shedding significantly decreased with the dose of AAV/meganuclease (OR=0.66; p=0.023, GLMM), and also with the duration of meganuclease therapy (OR=0.42; p<0.001, GLMM) in treated animals compared to controls. The data further indicate that overall, the quantity of virus shed (AUC) significantly decreased with the AAV/meganuclease dose at a rate of -0.36 log10 copy-days per 1012 increase in dose (LMM; p=0.028), and also with the duration of meganuclease therapy, at a rate of 0.48 log10 copy-days per additional week after treatment (LMM; p=0.017). No significant association was detected between the log10-transformed AUC and the interaction between time and dose (LMM; p=0.59).

The studies described above were performed using a triple AAV serotype/dual meganuclease approach, resulting in each animal receiving a total of 6 unique vectors (3 serotypes2 meganucleases). Clinical translation of such a complex regimen could raise manufacturing and quality control issues. We therefore sought to simplify AAV/meganuclease therapy to reduce the complexity of our therapeutic regimen. We took advantage of the dual cutting meganuclease m4, which recognizes a sequence in the duplicated gene ICP0 in the HSV-1 genome and was previously shown to induce significant decrease of latent viral loads in ganglia of latently infected mice8. Latently infected mice were administered a total dose of 51011 vg of either the combination of AAV9, AAV-Dj/8, and AAV-Rh10, or each single AAV serotype delivering the HSV1-specific meganuclease m4 (Fig.7a). Consistent with the results using the lower dose of 6 x 1011 of the triple AAV-dual MN therapy (Fig.4), ganglionic tissues from treated mice with the triple AAV combination delivering m4 showed a 73.9% (p<0.0001) and 43.7% (p=0.014) reduction in mean viral loads in SCG and TG, respectively, when compared to control untreated animals. When m4 was delivered using single AAV serotypes, the data confirmed that AAV9 on its own could recapitulate the viral load decrease seen with the triple AAV serotype combination, with 77.8% (p<0.0001) and 49% (p=0.0046) reduction in mean viral loads in SCG and TG, respectively (Fig.7b, c). Furthermore, mice having received AAV9 alone showed the lowest levels of liver inflammation of any of the groups, similar to those in the control liver (Fig.7d). At this reduced dose, regardless of the AAV serotype combination used, no detectable neurotoxicity was observed compared to the control animals (Fig.7e, f).

a Experimental timeline of ocular infection and meganuclease therapy. b, c HSV loads in SCGs (b; p<0.0001) and TGs (c; p=0.0046 for AAV9 and 0.0142 for 9-Dj/8-Rh10) from infected control and infected mice treated with m4 delivered by retro-orbital (RO)) injections of 51011 vg total of the single or triple combinations of AAV9, -Dj/8 and -Rh10. Percent decrease of HSV loads in treated mice (n=10 per group) compared to control mice (n=10) and statistical analysis (Ordinary one-way Anova, multiple comparisons with *p<0.05; **p<0.01, ****p<0.0001; ns: not significant). d Inflammatory cell foci (ICF) in liver sections from either HSV-infected control mice (n=10), or mice treated with m4 delivered using AAV single or triple combinations of AAV9, -Dj/8, and -Rh10 (n=10 per group); p=0.0009 for Rh10. e, f Severity scores of axonopathy (e) and inflammation (f) in TG from infected control mice (n=3 TG) and infected mice treated with m4 delivered using single or triple combinations of AAV9, -Dj/8 and -Rh10 (n=3 TG per group) and statistical analysis (Ordinary one-way Anova, multiple comparisons with ns: not significant; ***p<0.001). Each graph shows individual and mean values with standard deviation. The AAV viral loads are shown in Supplemental Fig.9jl. Source data are provided as a Source Data file.

To confirm that a simplified regimen composed of AAV9-m4 was also able to reduce peripheral virus shedding, latently infected mice were treated as above using AAV9 delivering either HSV1-specific meganuclease m4 or a catalytically inactive version (m4i) at a dose of 11012 vg. One month later, mice were subjected to two weekly rounds of JQ1 administration, followed by daily eye swabbing for 3 days (Fig.8a, b). A decrease of ganglionic viral loads of 89.6% (p<0.0001) and 69% (p=0.03) in SCG and TG respectively, was observed in m4-treated mice but not in mice treated with the inactive form of the meganuclease m4i (Fig.8c, d). Furthermore, 6 out of 9 control mice and 6 out of 10 m4i-treated mice shed virus after JQ1 reactivations, while only 3 out of 10 m4-treated mice had detectable virus shedding after reactivation (Fig.8ei). These data demonstrate that our simplified regimen can substantially reduce ganglionic viral loads, with an associated decrease in virus shedding after reactivation, and that these effects are dependent on an active enzyme and not on AAV itself. In this experiment, mice treated with m4 had slightly higher levels of liver ICF and TG axonopathy, but not more TG inflammation, compared to control mice (Fig.8jl).

a Experimental timeline of ocular HSV-1 infection, meganuclease treatment and viral reactivations with JQ1. b Schematic of active m4 and inactive m4i meganuclease. c-d, HSV loads in both SCGs (c; p<0.0001 for m4) and both TGs (d; p=0.003 for m4) from control infected mice and infected mice treated the active m4 or inactive m4i (n=10 per group). Percent decrease of HSV loads in treated mice compared to control mice and statistical analysis (unpaired one-tailed Mann-Whitney test with *p<0.05; ****p<0.0001; ns: not significant) are indicated. eg Virus titers in eye swabs collected at day 1 to 3 after each JQ1 reactivation from control infected mice (e) and infected mice treated with active m4 (f) or inactive m4i (g). h, i Area under the curve (AUC) analysis of virus shedding after first (h), and second (i) JQ1 reactivation from control infected mice and infected mice treated with active m4 or inactive m4i (n=10 per group). p values (unpaired one-tailed MannWhitney test) are indicated. j Inflammatory cell foci (ICF) in liver sections from either HSV-infected control mice, mice treated with active m4 or inactive m4i (n=10 per group); p=0.0234 for m4. k, l Severity scores of axonopathy (k; p=0.0007 for m4) and inflammation (l) in TG from HSV-infected control mice, mice treated with active m4 or inactive m4i (n=10 per group) with statistical analysis (Ordinary one-way Anova, multiple comparisons with ns: not significant; *p<0.05; ***p<0.001). Each graph shows individual and mean values with standard deviation. AAV viral loads are shown in Supplemental Fig.9mo. Source data are provided as a Source Data file.

Across our studies, we observed that ~10% of the animals treated with a high dose of AAV/meganuclease (231012 vg/animal, or approximately 691013 vg/kg) exhibited clinical signs consistent with hepatotoxicity, including weight change, bloating, and general health decline. When lower doses were evaluated, we observed substantially improved tolerability, both clinically and upon histopathological examination and ICF quantification. To further reduce hepatoxicity, we evaluated the use of neuron-specific promoters (Calmodulin Kinase II (CamKII) and human Synapsin (hSyn)) combined with the CMV enhancer17, to test the hypothesis that limiting enzyme expression to neuronal tissues would decrease or perhaps prevent liver toxicity (Fig.9a). We found that latently infected mice treated with a high dose (21012 vg) of AAV9-E/CamKII-m4 or AAV9-E/hSyn-m4 did not show any clinical signs of hepatotoxicity (weight change, general health decline, or ICF), in contrast to mice treated with 21012 vg AAV9-CBh-m4 (Fig.9b, c). Moreover, while liver inflammation increased over time in AAV9-CBh-m4-treated mice, it remained low in AAV9-E/CamKII-m4 (Fig.S11d). Intriguingly, histopathologic signs of neurotoxicity in TG from AAV9-E/CamKII-m4 or AAV9-E/hSyn-m4 treated mice were also similar to those in control mice, while they were significantly higher in TG from AAV9-CBh-m4 treated mice (Fig.9d, e). A decrease of ganglionic viral loads of 67.9% (p=0.07) and 70.4% (p=0.05) in SCG and TG respectively, was observed in AAV9-E/CamKII-m4-treated mice but not in mice treated with the AAV9-E/hSyn-m4 (Fig.9f, g). Assessment of m4 expression in neuronal tissues at different times post administration of either AAV9-CBh-m4 or AAV9-E/CamKII-m4 showed that the m4 expression increased over time but was in general slightly lower in tissues from AAV9-E/CamKII-m4-treated mice compared with AAV9-CBh-m4-treated mice (Fig.S11a, b). This may explain the slightly lesser degree of viral load reduction in AAV9- E/CamKII-m4-treated mice compared with AAV9- CBh-m4-treated mice (Fig.9f, g). We conclude that the AAV9-E/CamKII-m4 regimen retains efficacy and shows improved tolerability compared to AAV9-CBh-m4.

a Experimental timeline of ocular infection and meganuclease therapy. b Average weight change of infected control mice (n=13) or HSV-infected mice treated with m4 expressed from either the ubiquitous CBh promoter, or the neuronal promoters E/CamKII or E/hSyn (n=12 per group). c Inflammatory cell foci (ICF) in liver sections from either HSV-infected control mice (n=13), or mice treated with m4 expressed from either the CBh, E/CamKII or E/hSyn promoter (n=12 per group); p=0.00455 for CBh-m4. d, e Severity scores of inflammation (d; p=<0.0001 for CBh-m4) and axonopathy (e; p=<0.0001 for CBh-m4) in TG from infected control mice (n=10) and infected mice treated with m4 expressed from either the CBh, E/CamKII or E/hSyn promoter (n=12 per group) with statistical analysis (Ordinary one-way Anova, multiple comparisons with ns: not significant; *p<0.05; ****p<0.0001). f, g HSV loads in SCGs (f) and TGs (g) from infected control (n=10) and infected mice treated with m4 expressed from either the CBh, E/CamKII or E/hSyn promoter (n=12 per group). Percent decrease of HSV loads in treated mice compared to control mice and statistical analysis (Ordinary one-way Anova, multiple comparisons with p values; ns: not significant). Each graph shows individual and mean values with standard deviation. The AAV viral loads are shown in Supplemental Figure10ac. Source data are provided as a Source Data file.

Continued here:
Gene editing for latent herpes simplex virus infection reduces viral load and shedding in vivo - Nature.com

PE: Can you leverage lessons from the commercialization and distribution of rare products and apply to CGT?

Lattanzi: Most of the cell and gene therapies are rare and orphan products. Cencora has a long history in the specialty and rare and orphan space. 76% of our specialty distribution drugs actually have orphan drug designation. As weve built, our cell and gene offerings leveraged some of the experience from that specialty and rare and orphan space. You do have to make some adjustments because there are, as I mentioned earlier, some very unique needs for cell and gene. But there are also a lot of similarities. I know even as we develop new services and solutions or new operating procedures, were saying all the time this is actually not just good for cell and gene, but this is actually useful for other rare and orphan products as well.

Read more from the original source:
Leveraging Lessons from Commercialization and Distribution of Rare Products for Cell and Gene Therapy - Pharmaceutical Executive

The Latest

A recent Phase III open-label, single-arm study conducted by Pfizer in collaboration with Spark Therapeutics investigated the safety and efficacy of a gene therapy named Beqvez in adult male patients with moderately severe to severe hemophilia B. Spark Therapeutics was responsible for conducting all Phase 1 and 2 studies, whilePfizer assumed responsibility for pivotal studies, regulatory activities, and potential global commercialization. The clinical trial named the BENEGENE-2 study found that during follow-up, there were decreased annualized bleeding rates in patients when they received the gene therapy compared to when theywere treatedwith routine factor IX prophylaxis. The results showed that patientsgenerallytolerated Beqvez well, with no reports of death, serious adverse events, or thrombotic events.

Physicians Perspective

Hemophilia B is a rare genetic disorder that prevents normal blood clotting for people who cannot generate the factor IX protein, causing them to bleed more frequently and profusely. According to the World Federation of Hemophilia, 38,000 people are affected by this disorder. Patients with hemophilia B struggle with the commitment and lifestyle disruptions of regular (factor IX protein) infusions, spontaneous bleeding episodes, painful joint damage, and mobility issues. Traditionally, patientsneed toreceive routine infusions of Factor IX treatment to prevent and control bleeding. However, thiscurrentstandard treatment method does not alter the underlying disease process. With a gene therapy infusion, patients can receive a one-time treatment that helps them produce their own Factor IX. This alternative active source of endogenous Factor IX helps improve bleeding outcomes and reduces the need for Factor IX infusions. The single gene therapy injectionis expectedto be effective for at least10years.

Molecular Targets

Hemophilia Bis causedby mutations in the F9 genewhichcarries instructions for making Factor IX, a protein involved inthe formation ofblood clots. Beqvez delivers a highly functional version of F9 to liver cells, where blood clotting factorsare produced. The functional F9 gene is called Padua and encodes for a version of Factor IXthat has a5-10greaterclotting activity than the endogenously produced protein. The F9 geneis packagedinside a specialized adeno-associated virus, surrounded by an outer shell capsid thatis taken upby liver cells. Once the F9 gene componentis taken upby the liver cells, the endogenous machinery can begin to produce the enhanced version of the Factor IX protein.

Company History

Pfizer is an American pharmaceutical and biotechnology corporation headquartered in Manhattan, New York City. In addition to Beqvez, Pfizeralsohas gene therapies under investigation in phase 3 trials for hemophilia A and Duchenne muscular dystrophy. The companyis also testinganti-tissue factor pathway inhibitors to treat people with hemophilia A and B.

Further Reading: https://www.pfizer.com/news/press-release/press-release-detail/us-fda-approves-pfizers-beqveztm-fidanacogene-elaparvovec

2024 2 Minute Medicine, Inc. All rights reserved. No works may be reproduced without expressed written consent from 2 Minute Medicine, Inc. Inquire about licensing here. No article should be construed as medical advice and is not intended as such by the authors or by 2 Minute Medicine, Inc.

Originally posted here:
Pfizer: Beqvez Gene Therapy Decreases Bleeding Rates in Hemophilia B Patients - 2 Minute Medicine

The result comes from a phase 1/2 trial of the biotechs experimental adeno-associated virus (AAV) gene therapy DB-OTO.

Opal Sandy, now aged 18 months, was born with profound deafness attributed to auditory neuropathy, a condition stemming from disrupted nerve impulses between the inner ear and the brain.

One of the youngest children in the world to receive a gene therapy for genetic deafness, she was treated at 11 months of age.

A second child in the CHORD trial, treated at age four, has also seen positive results after six weeks, according to data presented at the American Society of Gene and Cell Therapy (ASGCT) annual conference.

Professor Manohar Bance, an ear surgeon at Cambridge University Hospitals NHS Foundation Trust and the lead researcher for the trial, conveyed to the PA news agency that the findings surpassed his initial hopes and expectations, potentially indicating a breakthrough in treating patients with this form of hearing impairment.

We have results from [Opal] which are very spectacular so close to normal hearing restoration. So we do hope it could be a potential cure, he said.

Regeneron's DB-OTO aims to address mutations found in the otoferlin gene, known as OTOF, responsible for encoding a substantial protein present in hair cells within the cochlea, which plays a crucial role in converting sound into neuronal signals.

In the ongoing study, patients receive a single intracochlear injection of DB-OTO in one ear and a cochlear implant in the other.

Following treatment, the children then take part in several hearing and brain tests, focusing on the tones needed for speech.

The trials secondary endpoints include hearing improvement, measured using auditory brainstem response (ABR) and behavioural audiometry with pure-tone audiometry (PTA).

The opportunity of providing the full complexity and spectrum of sound in children born with profound genetic deafness is a phenomenon I did not expect to see in my lifetime, said Lawrence Lustig, trial investigator and otolaryngologist at Columbia University.

These impressive results showcase the revolutionary promise of DB-OTO as a potential treatment for otoferlin-related deafness, and we are excited to see how this translates into an individuals development, especially since early intervention is associated with better outcomes for speech development.

"With the DB-OTO CHORD trial now enrolling participants in sites across the U.S. and Europe, were part of the beginning of a new era of gene therapy research that looks to create treatment options that address the root cause of profound genetic deafness.

At the start of the study, both pediatric patients showed no reaction to sounds delivered at maximum levels of 100 decibels or more, and there was no evidence of a signal reaching the hearing centres of the brain using electrophysiological auditory evoked response (ABR) testing.

However, at 24 weeks Opal observed improvement of her hearing to normal levels, with an average 80 decibels improvement from baseline.

The second child observed an average 16 decibels improvement in hearing response, compared to baseline, at six weeks.

In addition, the surgical procedure and the gene therapy were found to be well tolerated, with no treatment-related adverse events or serious adverse events.

According to Regeneron, the study is still recruiting participants across the US and Europe and aims to enrol approximately 22 children with otoferlin hearing loss aged 17 years and younger.

View original post here:
Regeneron gene therapy restores hearing in profoundly deaf child - BioPharma-Reporter.com

The two posters demonstrate in vivo superiority of circular RNA vs. linear mRNA-based expression.

Circio has generated results demonstrating that the circVec 2.1 design performs very well in vitro. We have now confirmed this in vivo with statistically significant higher expression level and durability for circVec 2.1 DNA vectors compared to standard linear mRNA-based expression, said Dr Thomas Hansen, chief technology officer at Circio.

These results provide an important technical proof-of-concept for Circios technology platform in an animal model. We now have confirmation for our expectation that this could translate into improved gene therapies for patients in the future.

In recent experiments, Circio has observed up to four months circVec durability in vivo. This substantially outperforms mRNA vector expression. Following these results, we can rapidly advance to design and test circVec in several AAV and DNA-based vectors. This will validate these very promising data in therapeutically relevant formats.

At ASGCT, Circio also presented its dual-function remove and replace concept for Alpha-1-antitrypsin deficiency (AATD).

AATD is a genetic disease that causes severe symptoms in the lung and liver.

There are currently no robust therapeutic options available for this indication and the condition still represents a major unmet medical need, with over 200,000 patients affected in the USA and EU alone.

With the technologically differentiated circVec remove-&-replace format, Circio has developed a unique gene therapy concept that it claims can deal with both the lung and liver-associated symptoms in one single therapeutic.

AATD is a challenging genetic disease to treat. This is in part due to the two distinct pathologies in the liver and lung. We have now established and technically validated circVec constructs that can both replenish functional wild-type AAT and specifically remove more than 90% of the mutated protein, said Dr. Victor Levitsky, chief scientific officer at Circio.

This is challenging to achieve because the functional and mutant forms are very similar. By using circular RNA-based AAT expression, Circio is uniquely able to separate the two species for mutant-specific knockdown, thereby solving two problems with one single product.

Continued here:
Circio debuts proof-of-concept for circVec gene therapy at ASGCT 2024 - BioPharma-Reporter.com

EBT-101, a CRISPR-based gene-editing therapy from Excision BioTherapeutics, was safe and well tolerated in a Phase I/II study, but it did not prevent viral rebound in the first three participants who stopped antiretroviral treatment, according to a presentation last week at the American Society of Gene & Cell Therapy (ASGCT) annual meeting.

Excision put a positive spin on the findings, noting that favorable safety data is a necessary step on the path to developing therapies for latent viral infections. The company also touted promising early results from studies of CRISPR-based therapies for herpes simplex virus and hepatitis B. But the HIV rebound news is disappointing, and it underscores the importance of remaining wary of exaggerated claims from industry and the mainstream press about the state of HIV cure research.

Initial data from the EBT-101-001 trial provides important clinical evidence thatagene editingtreatment modality can be safely delivered fortargetingthe HIV DNA reservoirs in human cells, study investigator Rachel Presti, MD, PhD, of Washington University St. Louis School of Medicine, said in a news release. This study provides researchers with invaluable insights for how CRISPR technology can be applied for addressing infectious disease and was an important first step towards additional programs designed to optimize this treatment modality for treating the millions of individuals who are impacted by HIV and other infectious disease.

Antiretroviral therapy can keep HIV replication suppressed indefinitely, but the virus inserts its genetic blueprints into the DNA of human cells and establishes a long-lasting reservoir that the drugs cant reach. This integrated HIV DNA lies dormant in resting T cells during treatment, but it can start churning out new virus when antiretrovirals are stopped, making a cure nearly impossible. The only way to tell whether an intervention leads to long-term remission is to discontinue antiretroviral therapy with careful monitoring, known as an analytic treatment interruption.

Kamel Khalili, PhD, of Temple University, and colleagues have been studying gene therapy to cure HIV for more than a decade. Their work employs CRISPR-Cas9, a technology that combines guide RNAs that home in on specific segments of DNA and a nuclease enzyme that cuts the genetic material at the desired site. In 2014, they reported that a CRISPR-Cas9 tool could cut out a segment of integrated HIV DNA necessary for viral replication in a laboratory study. Another study published in 2019 showed that this approach could remove integrated HIV genes and clear latent viral reservoirs in mice.

This led to the development of EBT-101, a CRISPR-based therapy delivered by an adeno-associated virus that uses dual guide RNAs to target three sites on the integrated HIV genome. Making cuts at these locations prevents the production of intact virus. Last August, researchers reported that a single dose of a simian version of the therapy safely and effectively removed an HIV-like virus from viral reservoirs in monkeys on antiretroviral therapy, but this study did not include a treatment interruption.

The first human clinical trial of EBT-101 (NCT05144386) started in 2022, enrolling people on antiretroviral therapy with a stable undetectable viral load. The study protocol called for participants who maintained viral suppression at 12 weeks after receiving the gene therapy to undergo an analytic treatment interruption.

At the European Societyof Gene & Cell Therapy annual meeting last October, Presti reported that EBT-101 was detectable in the blood of the first three treated participants after a single IV infusion at the initial dose level. EBT-101 was well tolerated with only mild temporary side effects. She did not present treatment interruption outcomes, but that didnt stop the Daily Mail from proclaiming that a cure for HIV could be months away.

Presti gave an update last week, and the news generally wasnt good. Of the five participants who have so far received the initial dose of EBT-101, three stopped antiretroviral therapy. Unfortunately, all three experienced viral rebound and had to restart their antiretrovirals. This likely occurred because the gene therapy did not reach all cells harboring latent HIV, and even a very small number of cells containing residual HIV DNA is enough to reignite viral replication.

But the news was not all bad. One EBT-101 recipient was able to maintain viral suppression for four months after treatment discontinuationconsiderably longer than it typically takes for the virus to rebound after stopping antiretrovirals. This suggests that EBT-101 or similar CRISPR therapies might play a role in a combination functional cure strategy.

We know that many people were hopeful that a first trial could provide evidence of a possible cure for HIV because the field has been waiting over 20 years for a cure, Excision senior vice president William Kennedy, MD, said in a news release. However, it was essential that this clinical trial establish safety for EBT-101 as a gene therapy product as well as safety related to the use of CRISPR for the field.

Excision is now testing a higher dose of EBT-101 in a second cohort and is exploring new CRISPR delivery methods that might be more efficient than the adeno-associated virus vector. One possibility is lipid nanoparticles like the ones used to deliver messenger RNA (mRNA) in COVID-19 vaccines.

The company is also exploring CRISPR-based approaches for other latent viral infections. Herpes simplex virus (HSV) persists in nerve cells, and it can reactivate to cause cold sores, genital herpes or eye inflammation (keratitis). Hepatitis B virus (HBV) establishes chronic infection in the liver, where it can potentially lead to cirrhosis and liver cancer. Unlike HIV and other retroviruses, however, HSV and HBV do not integrate their genetic blueprints into the chromosomes of human cells, so they may be easier to remove.

In other presentations at the ASGCT meeting, researchers reported preclinical results for another experimental CRISPR therapy dubbed EBT-104, showing that a single dose reduced HSV DNA by more than 99% in laboratory cell cultures. Whats more, it eliminated viral shedding in 11 of 12 rabbits with herpes keratitis, according to the news release.

In other preclinical research, a single dose of EBT-107a CRISPR compound delivered by lipid nanoparticlesreduced HBV DNA, hepatitis B surface antigen and hepatitis B e antigen by 98%, 97% and 92%, respectively, in a mouse model of hepatitis B. Unlike CRISPR delivered by viral vectors, EBT-107 in nanoparticles and could potentially be given as multiple doses to reach more latent virus.

Click here for more news about HIV cure research.

More:
CRISPR HIV Gene Therapy Disappoints in Early Study - POZ

Gene therapy research is booming. Since the U.S. Food and Drug Administration (FDA) issued its first approval for a gene therapy in 2017,1 oncology researchers have been breaking barriers in gene therapy trials, followed by an explosion in mRNA research during the COVID pandemic. Today, this trailblazing science is providing new ways to approach rare diseases and new hope when other investigational interventions have failed. In fact, the majority2 of approved gene therapies are for rare diseases14 are currently in Phase III trials for 10 rare diseases and 45 gene therapies are in early stages of development to treat 30 rare diseases.

We see great potential for gene therapies, said Leslie Johnston, senior vice president of biotech delivery for Parexel. As more products are approved, it will gain traction and more companies will look to expand their therapies into other therapeutic indications. This progress presents tremendous potential to change more patients lives across many different diseases.

This could be gene therapys moment. But to fully seize it, the industry must clear some complex hurdles. Gene therapies pose several unique challenges for clinical research, including ethical and safety considerations, regulatory hurdles, precarious logistics, and potentially staggering costs. These challenges may already be having ramifications: New U.S. patients treated with gene therapies approved or in development are expected to fall by one-third from 2025 to 2034.3

The key to clearing these hurdles? Cooperation between sponsors, sites, regulators, patients, and other stakeholders is essential to expediting the advancement of life-saving gene therapies.

However, obtaining regulatory approval under these conditions is time consuming and resource intensive. To avoid hampering scientific progress, regulators should aim to ensure that requirements are appropriately rigorous without being unmanageably onerous.

Thankfully, the FDA is paying close attention to gene therapy and has demonstrated a desire to work with drug developers toward the success and approval of these treatments. Dr. Peter Marks, Director of the Center for Biologics Evaluation and Research (CBER) at the FDA, has expressed his hope for an exponential, if not logarithmic, increase in gene therapy approvals.

There is a lot of excitement that this could potentially make a big difference for the treatment of human disease, said Dr. Marks in his remarks to the National Press Forum last November.4

The FDA is going beyond mere rhapsodizing and taking action to accelerate gene therapy. Last year, the agency launched a pilot program called Support for Clinical Trials Advancing Rare Disease Therapeutics, or START.5 This program is designed to accelerate the development and approval process for treatments targeting rare diseases by providing regulatory guidance, assistance, and incentives to sponsors conducting clinical trials in this field. The program represents an important step forward in fostering innovation and collaboration between regulatory bodies and sponsors.

In addition, the FDA is working to harmonize global requirements for the review of gene therapies.6 Encouraging and facilitating international cooperation and harmonization of regulatory standardsincluding mutual recognition agreements and shared regulatory pathways for multinational clinical trialscan help streamline gene therapy development globally and help bring innovations to patients faster.

Even with this progress, regulators should continue to help accelerate gene therapy research by streamlining regulatory pathways specifically tailored to gene therapies. This means providing clear guidance on requirements for preclinical and clinical development, fostering collaboration between stakeholders to share knowledge and best practices, and offering expedited review processes for gene therapy products aimed at treating serious or life-threatening diseases.

With a staggering 2,500 cell and gene therapy investigational new drug applications (INDs) on file, the FDA approved just five cell and gene therapies in 2023. Dr. Marks has suggested that accelerated approval, which has successfully advanced cancer and HIV/AIDS treatments, may be the most appropriate path for this new category of treatments. But regulators also need to commit to proactively partner with developers to understand the patient population and the risks and benefits of each new therapy.

Likewise, researchers, industry stakeholders, and patient advocacy groups should engage with regulators to help them understand the unique challenges and opportunities in the field of gene therapy. This can help regulators adapt regulatory frameworks to ensure patient safety while expediting the development and approval of promising treatments.

By working closely with clinicians and regulators, sponsors can ensure that the trial development process aligns with clinical needs and regulatory standards. Sponsors should have a thorough understanding of FDA requirements pertaining to design, preclinical testing, and long-term follow-up. Better alignment from the outset will lead to more efficient trial designs, faster regulatory approvals, and ultimately quicker patient access to therapies.

For example, sponsors working with mRNA and other genetically engineered therapies in North America not only have to go through institutional review board (IRB) review, they also have to navigate additional requirements from the U.S. National Institutes of Health (NIH) Office of Science Policy Guidelines for Research Involving Recombinant or Synthetic Nucleic Acid Molecules (NIH Guidelines).7 These requirements usually involve an additional biosafety risk assessment review from an institutional biosafety committee (IBC) in addition to IRB review.

NIH Guidelines apply for any research involving recombinant or synthetic nucleic acids (e.g. genetically engineered materials) that receives NIH support or takes place at sites that have received NIH support for such research. Even when there is zero NIH support, IBC review is considered a best practice. IBC review and inspection helps sites ensure they are fully prepared by identifying areas for improved biosafety protections and calling out gaps in current standard operating procedures (SOPs). Proactive coordination and integration of these separate review processes can speed trial timelines and help sponsors consistently address any potential concerns or issues.

Sites can also be better prepared by pre-registering an IBC. The NIH takes six to eight weeks or more to approve a new registration, in addition to IBC review timeso by registering an IBC before they even have a trial, sites can save a month or more in startup time over a site that waited to register.

Clinical trial sites looking to host gene therapy studies must be prepared in other ways, as well, both in terms of knowledge and infrastructure. Gene therapy studies require specialized infrastructure for manufacturing, storing, and administering genetic material to adhere to strict biosafety guidelines. Something as simple as having an upholstered chair in the infusion roomwhich would pose an unacceptable contamination risk if genetic materials were to spillwould require the site to rethink their current processes. Rigorous training is also key due to the added risk of spreading genetic material to caregivers and others in close contact with patients. Research staff must be specially trained to handle, deliver, and dispose of this material safely.

Of course, these measures can seem intimidating for sites that are already cost-constrained. Large academic medical centers with more resources and experience are more likely to be well-positioned for these studies. For instance, they may already have conducted bench, animal, and/or agricultural research with genetic engineering or have the funding to make any needed adjustments such as purchasing special equipment. But to maximize the potential number of sites where this research can be conductedand therefore reach more potential participantssponsors might consider providing help in the form of financial assistance, training curricula, SOP guidance, and more to smaller sites seeking to conduct gene therapy research.

Logistical complexities, depending on the investigational medicine and therapeutic area, are among the most complicated challenges in gene therapy trials, added Johnston. From collecting the specimen from the patient, modifying it, storing it, transporting it, and then returning it back to the patient all comes with tremendously unique logistical challenges and requires equally unique equipment, technology, and expertise. And it can be cost-prohibitive.

First, its crucial for patients to adhere strictly to the protocol provided by the clinical trial team, including following medication schedules, maintaining specific hygiene practices, and attending all study visits. They should strive to maintain optimal health to enhance the bodys response to gene therapy. And to avoid delays, patients should maintain open and honest communication with the clinical trial team, reporting any changes in symptoms, side effects, or general health as soon as they occur.

Trial participants also need to be in it for the long haul. Because gene therapy interventions aim to produce lasting effects, even cures, they typically require long-term patient follow-up to assess efficacy and safety. But they may also need to have incredible patience.

Johnston explained, There are many complexities that can impact study progress. For example, unpredictable logistical challenges like a weather event or vehicle accident could delay a temperature-sensitive delivery to a site, or data review outcomes could require an indeterminate pause period. Patience and agility are must-haves, but it is difficult for patients potentially depending on this new therapy to save or change their lives.

Lastly, the industry cannot forget the patient. Involving patients and patient advocacy groups in the regulatory process can help ensure that the development of gene therapies is aligned with patient needs and priorities, as well as shed light on risk-benefit perspectives from a patients viewpoint. The more these perspectives are considered from the beginning, the greater the chance of a trials success.

Rita Naman, co-founder of the Mighty Milo Foundation, emphasizes the need for a more collaborative and patient-centered approach to gene therapy development.

For ultra-rare diseases like SPAX5, gene therapy offers a glimmer of hope where traditional treatments do not. But logistical hurdles make these therapies expensive and inaccessible, explained Naman. Closer collaboration with patients, industry, and regulators could streamline these processes, drive costs down, and speed trials. Patients like my son, and their caregivers, plus advocacy groups should be invited into the earliest discussions to prevent false starts or missed milestones in gene therapy development especially as the patients priorities dont always line up with the sponsors.

Patients want access to novel gene treatments, and they want it fast. Sponsors want to deliver but fight logistical and financial obstacles. Regulators want to ensure safety first, especially considering such new, promising science, concluded Johnston. These three goals may seem conflicting at times, so we need to strike a balance of safety and speed, so patients dont miss their only potential treatment opportunity.

References 1. https://www.fda.gov/news-events/press-announcements/fda-approval-brings-first-gene-therapy-united-states 2. https://www.oaepublish.com/articles/rdodj.2023.29 3. https://www.bloomberg.com/news/articles/2024-02-20/gene-therapy-makers-struggle-to-find-patients-for-miracle-cures?embedded-checkout=true 4. https://nationalpress.org/topic/peter-marks-fda-crispr-sickle-cell-gene-therapy/ 5. https://www.ajmc.com/view/new-fda-pilot-program-will-provide-hands-on-regulatory-guidance-for-cell-and-gene-therapy-trials-in-rare-disease 6. https://endpts.com/fda-develops-pilot-to-harmonize-international-gene-therapy-regulations/ 7. https://osp.od.nih.gov/wp-content/uploads/NIH_Guidelines.pdf

See more here:
The Transformative Potential Of Gene Therapy - Contract Pharma