Genetic Engineering in the Press by GEG

T lymphocytes possess a retinoic acid receptor alpha (RARα) that is known to control gene expression programs in the nucleus, but now also appears to function outside the cell nucleus to coordinate the initial events triggered at the cell surface that lead to T cell activation. The role of RARα has been revealed thanks to the development of CRISPR techniques, advances in imaging and mass spectrometry. The aim of this research is to identify a new pathway, or set of pathways, that could be exploited to control autoimmune diseases and inflammation, or boost immunity to eradicate tumors or fight infections. Researchers had published research showing that retinoic acid (RA) triggers nuclear RARα and the expression of genes important for the differentiation of regulatory T cells. Scientists also knew that RA, present in the blood and taken up by T cells, is then transported to the nucleus by a molecule called cellular retinoic acid-binding protein 2 (CRABP2). CRABP2 in the cytoplasm binds to RA and transports it into the cell nucleus, where it activates nuclear RARα. The researchers set out to understand which other RARα proteins interact with it. This work revealed interactions with the ZAP70 kinase. For T cells, phosphorylation prompts key proteins to act when a threat is near. 

The European Commission has approved axicabtagene ciloleucel (axi-cel; Yescarta) for the treatment of adult patients with diffuse large B-cell lymphoma (DLBCL) or high-grade B-cell lymphoma (HGBL) who relapse within 12 months of completing, or are refractory to, first-line chemoimmunotherapy. The approval is based on the results of the pivotal Phase 3 ZUMA-7 trial (NCT03391466) which is a global, multicenter, open-label, randomized Phase 3 study evaluating the safety and efficacy of a single infusion of axi-cel vs SOC. This study consists of platinum-based salvage chemotherapy followed by high-dose chemotherapy and stem cell autotransplantation in patients with relapsed/refractory DLBCL within 12 months of first-line therapy. At a median follow-up of 2 years, axi-cel led to a significant improvement in the primary end point of event-free survival (EFS ) compared with standard of care (SOC; HR, 0.40; 95% CI, 0.31-0.51; P < 0.001). Median EFS was 8.3 months (95% CI, 4.5-15.8) versus 2.0 months (95% CI, 1.6-2.8) with axi-cel and SOC, respectively. The 2-year SES rate was 41% (95% CI, 33%-48%) with axi-cel versus 16% (95% CI, 11%-22%) with SOC. Axi-cel is now the first CAR T-cell therapy approved for patients in Europe who do not respond to first-line therapy.

Researchers in China show that a technique used to replace diseased mitochondria does not affect early development.

The CAR T-cell therapies axicabtagene ciloleucel (axi-cel; Yescarta), tisagenlecleucel (Kymriah) and lisocabtagene maraleucel (liso-cel; Breyanzi) are all FDA-approved for the treatment of patients with DLBCL. However, patients with primary and secondary CNS lymphoma have been excluded from clinical trials using these agents due to the risk of toxicities such as CRS and ICANS. According to a systematic review and analysis of published data presented at the 2022 Transplantation and Cell Therapy Meetings, Axicabtagene ciloleucel, Tisagenlecleucel and liso-cel elicited responses without increased risk of cytokine release syndrome or effector immune cell-associated neurotoxicity in patients with primary or secondary central nervous system (CNS) large B-cell lymphoma (LBCL). In the study, 47.6% of the patients evaluated were male. In addition, 28.3% of patients had primary CNS lymphoma and 71.8% of patients had secondary CNS lymphoma. Of these patients, 53.5% achieved CR, 14.1% had PR and 36.6% had progressive disease. The PFS rates at 1 month, 3 months, 6 months and 12 months were 72.1%, 57%, 44.2% and 37.5%, respectively. 

 What is the overview of CRISPR clinical trials in rare diseases? The Phase 1 trial of NTLA-2001, a CRISPR-Cas9 therapeutic candidate, was the first CRISPR therapy to be injected directly into humans via the bloodstream. It is designed to treat hereditary transthyretin amyloidosis (HTRA) by reducing the levels of mutated transthyretin (TTR) protein in the blood by inactivating the TTR gene in liver cells. The U.S. FDA granted orphan drug designation to NTLA-2001 for TTR in October 2021. EDIT-101 is a CRISR-Cas9-based gene editing candidate designed to correct mutations in the CEP290 gene for Leber Congenital Amaurosis 10 (LCA10), which accounts for nearly one-third of all LCA cases. While CEP290 mutations disable light-sensing cells in the retina, the retinal cells themselves are still present and viable in LCA10, giving hope that EDIT-101 may improve vision in this patient group. The NTLA-2002 clinical trial for its CRISPR therapy for HAE was approved in New Zealand in October. NTLA-2002 is delivered as Cas9 mRNA and gRNA via lipid nanoparticles, and preclinical studies have demonstrated a sustained and therapeutically relevant reduction in plasma kallikrein activity after a single dose in non-human primates. 

Silvia Corvera and Michael Czech have spent decades researching type 2 diabetes, trying to find new therapies for the metabolic disease that affects up to 10% of the population. Recently, they have seen promising results for a CRISPR-based cell therapy in mice, and clinical trials are the longer-term goal thanks to smart use of the gene-editing technology that reduces the risk of adverse side effects. The new therapy is based on the implantation of fat tissue that has been genetically engineered to burn fat and glucose and secrete various factors that regulate metabolic health. The researchers realized that when a gene called Nrip1 was turned off in fat cells, they responded much better to insulin. Over time, it became clear that Nrip1 suppresses insulin sensitivity and many genes usually expressed in brown fat cells, including Ucp1, which causes them to burn fat. If modified cells were then transplanted into mice, their ability to burn fat and glucose and secrete metabolic factors could have beneficial effects on metabolic health.  

Results from early clinical trials suggest that CAR T cells can be successfully adapted to target brain tumors. A Phase 1 analysis of a next-generation CAR T cell therapy has shown that it can target and kill tumor cells while eliciting an immune system response to aid in this effort. An ongoing trial is evaluating an IL13R-alpha-2-directed CAR-T administered locoregionally for the treatment of adults with relapsed or refractory glioblastoma. Patients receive at least three infusions of CAR-T cells administered intracranially. Initial results have shown that weekly CAR T cell dosing is safe at all dose levels studied, with no dose-limiting toxicity or flu-like symptoms. Data from a cohort of patients whose treatment was optimized for dose level and manufacturing process showed a median SG of 10.5 months in heavily pretreated patients whose disease progressed after prior treatment. By comparison, historical data showed an approximate median SG of 5 months to 9 months for patients with recurrent glioblastoma. His group is now analyzing why some patients have achieved a particularly good response to CAR T cells.

B-cell acute lymphoblastic leukemia (B-cell ALL), the most common type of cancer in children, is a type of blood and bone marrow cancer that develops in early forms of B cells - white blood cells - and progresses rapidly if left untreated. A retrospective study by Northwestern Medicine researchers found that tisagenlecleucel, a CAR T-cell immunotherapy drug, demonstrated safety and efficacy in pediatric patients with relapsed and refractory B-cell ALL, even when the drug did not meet the FDA's strict manufacturing standards. The findings, published in Blood, may help expand therapeutic interventions for these patients who have otherwise exhausted all other treatment options, according to Jenna Rossoff, MD, instructor of pediatrics in the Division of Hematology, Oncology and Stem Cell Transplantation and lead author of the study.

Mosquitoes spread viruses that cause potentially deadly diseases such as yellow fever. New research funded by the U.S. military is using CRISPR-Cas 9 to target specific fertility-related genes in male mosquitoes to make them infertile and slow the spread of disease.

Using gene editing in male Aedes aegypti, the researchers found that the mutant male mosquitoes produced no sperm, and unlike previous studies, the sterile studs were otherwise perfectly healthy. However, the team did not know whether the defective sperm from the sterile males was necessary to render the female mosquitoes infertile, or whether the transfer of seminal fluid was sufficient. In addition, to manage populations, scientists use a vector control practice called the sterile insect technique in which they rear large numbers of sterile male insects and then release these males in numbers that exceed their wild counterparts. Females that mate with sterile males before finding a fertile one are themselves rendered infertile, thus decreasing the size of the next generation.

The development of CAR T cells is one of the most important recent advances in the field of immunotherapies targeting cancer. This treatment is mainly used for patients suffering from leukemia or lymphoma. However, the application of CAR T cell therapy to solid tumors is still challenging. For many cancers, there is an abundance of specific proteins on the surface of the tumor that most often exist on the surface of healthy cells. CAR T cells therapy is not able to distinguish between them, which can lead to fatal side effects.

However, American and Finnish researchers have identified a new way to program CAR T cells to kill only cancer cells. The researchers have developed a new CAR T cells engineering technique based on a two-step process of identifying HER2-positive cells and the antigen density. HER2 is a characteristic protein of breast, ovarian and abdominal cancer, but is also found in large numbers on the surface of tumor cells. One of the researchers' goals is to use this technology against a large wide of solid tumors.

A new study has revealed that a new T cell genetically modified by researchers at the University of Arizona is capable of targeting and attacking the pathogenic T cells responsible for type 1 diabetes, which could lead to new immunotherapy treatments. Michael Kuhns, PhD, Associate Professor at the University Arizona College of Medicine - Department of Immunobiology in Tucson, has copied the evolutionary design of killer T cells to design with a five-module chimeric antigen receptor, 5MCAR. Dr. Kuhns worked with Thomas Serwold, PhD, of the Joslin Diabetes Center, affiliated with Harvard Medical School, to test the 5MCAR T cells in a non-obese diabetic mouse model with promising results. The results were recently published in the proceedings of the National Academy of Sciences. They therefore designed a 5MCAR that would direct killer T cells to the autoimmune T cells that are involved in type 1 diabetes. The 5MCAR T cells would completely eliminate the harmful T cells that invade the pancreas. Drs. Kuhns and Serwold recently received a bridging grant from the National Institute of Allergy and Infectious Diseases to continue their research on the use of 5MCAR T cells to prevent autoimmune diseases. 

The use of BCMA D8 Fab CAR-directed CAR T cell therapy and AUTO8 dual-targeted CAR T cell therapy is safe and feasible in patients with relapsed and refractory multiple myeloma, according to initial results from the ongoing Phase 1 MCARTY trial (NCT04795882). After a median follow-up of 6 months, the objective response rate was 100% in a group of 11 patients who received an infusion. Three patients had a partial response, a very good partial response and a complete response or strict complete response. Overall, progression-free survival was not achieved. In addition, the investigators reported no cases of immune effector cell-associated neurotoxicity (ICAN) syndrome. The investigators aimed to develop a CAR ultra-sensitive to B-cell maturation antigen (BCMA) that co-targets CD19 when using a CAR, as well as to test the product in a clinical study. 

Many ovarian tumors contain mesothelin. Researchers wanted to test three types of CAR molecules programmed to attack this particular protein. They repeatedly exposed ovarian cancer cells to CAR T cells in test tubes and conducted several experiments in mice. All three types of CAR T cells significantly prolonged the lives of the mice with cancer compared to those in the control group. Mice that were injected with T cells expressing this particular molecule saw a reduction in tumor size and lived even longer than the others. Several of the mice were even cured. The researchers hope that this discovery will pave the way for a clinical study. Their goal is to predict the optimal conditions for producing CAR T cells that can infiltrate and attack the tumor and survive in the bodies of women with ovarian cancer. 

Inherited heart muscle diseases can cause sudden heart failure or progressive heart failure in young people. It is estimated that one in 250 people worldwide is affected by genetic cardiomyopathies, with a 50/50 chance that they will pass their defective genes on to each of their children. In many cases, several members of a family will develop heart failure, require heart transplants, or die suddenly of cardiac causes at a young age. However, an injectable cure for inherited heart muscle diseases that can kill young people in the prime of their lives could be available within a few years, thanks to the team of researchers who won the British Heart Foundation's Big Beat Challenge. The prize money is £30 million. The team will take the revolutionary CRISPR gene-editing technology to the next level by deploying ultra-precise techniques, called basic and prime editing, in the heart for the first time. These revolutionary approaches use ingenious molecules that act like tiny pencils to rewrite unique mutations that are buried in the DNA of heart cells in people with genetic cardiomyopathies.

GC012F is an autologous, genetically engineered, bispecific CAR-T that targets B-cell maturation antigen and CD19 proteins on the surface of cancer cells. The new cell therapy, which has previously received FDA orphan drug designation, is manufactured within 24 hours of apheresis using Gracell's proprietary FasT CAR platform. A single dose of an investigational chimeric antigen receptor T-cell therapy induced an objective response in up to 90% of patients with relapsed or refractory multiple myeloma, results from the Phase 1 study showed. Data from the trial, presented at the ASCO Annual Meeting, indicated that the agent had a manageable safety profile. Twenty-five (89.3%) of the 28 patients achieved objective treatment responses after a single infusion of GC0-12F, including 21 (75%) with a negative complete response to minimal residual disease or a strict complete response to treatment. Twenty-four (86%) patients had a very good partial response or better. Researchers reported few cases of high-grade cytokine release syndrome and no treatment-related neurotoxicity.   

The FDA has granted accelerated designation to PRGN-3006 for the treatment of adults with relapsed or refractory acute myeloid leukemia. PRGN-3006 (Precigen Inc.) is a CAR T cell therapy that targets the CD33 protein on the surface of cancer cells. This investigational therapy, designed to reduce manufacturing time to 1 day, is based on Precigen's UltraCAR-T therapeutic platform. The therapy uses the Sleeping Beauty non-viral gene editing system to simultaneously express CD33-targeted CAR, membrane-bound interleukin-15 and a therapeutic "kill switch" that conditionally eliminates CAR T cells for increased safety. PRGN-3006 is being evaluated in a single-center Phase 1/phase 1B dose escalation/extension study to determine safety and recommended Phase 2 dose for adults with relapsed or refractory AML, high-risk myelodysplastic syndrome or chronic myelomonocytic leukemia. Study participants will receive a single IV dose of the treatment with or without prior lymphodepletion. Interim results from the study, presented last year at the ASH Annual Meeting, showed an overall response rate of 50% in the first six patients in the lymphodepletion cohort. Safety results showed no dose-limiting toxicities. One of the 15 patients developed a grade 3 cytokine release syndrome. 

Memorial Sloan Kettering Cancer Center has paused a phase 1 clinical trial of ATA2271, a chimeric antigen receptor T-cell therapy, after the death of a patient treated with the experimental therapy. ATA2271 (Atara Biotherapeutics) is an autologous CAR-T cell therapy that targets the mesothelin antigen expressed on the surface of certain solid tumors. It is being evaluated in a single-center, dose-escalation Phase 1 study in patients with malignant pleural mesothelioma. Six patients in the study received one of the first two dose levels without any dose-limiting toxicities. The deceased patient had a history of comorbidities and other malignancies and was the first to receive treatment at a dose of 6 × 10 6 cells/kg.  

In 2018, biophysicist He Jiankui shocked the world when he announced that he had used the CRISPR genome editing technique to modify implanted embryos and led to the birth of two children. The genomes of the girls were modified to induce HIV resistance. Just after the egg was fertilized, the scientists injected CRISPR Cas 9 reagents into the fertilized egg. They wanted to make targeted genetic modifications, the goal was that this would lead to people who would be resistant or even immune to HIV, the virus that causes AIDS. The genetic modification was supposed to deactivate a protein that acts as a kind of molecular door handle for HIV. According to Nature Biotechnology, his toddlers are now healthy. Researchers are investigating how the changes to the girls' genomes might result in health benefits or risks, how their condition might be monitored, given He's imprisonment and the closure of his lab, and how other researchers might ethically study the genetically altered girls.

On October 1, 2021, the FDA approved brexucabtagene autoleucel for patients with relapsed/refractory B-cell precursor ALL. The approval was based on results from the phase 1/2 ZUMA-3 trial (NCT02614066), which showed a complete response/complete response with incomplete blood count recovery (CR/RCi) rate of 70.9%, which included a 56.4% CR rate. At the data cutoff date, 31% of patients who achieved CR/RCi were in continuous remission without subsequent allogeneic stem cell transplantation. The median duration of response with censoring on subsequent allogeneic stem cell transplantation, relapse-free survival, and overall survival (OS) was 12.8 months (95% CI, 8.7-not estimable [NE]), 11.6 months (95% CI, 2.7 -15.5), and 18.2 months (95% CI, 15.9-NE), respectively. Median overall survival was not achieved in patients who achieved CR/RCi. These data are unprecedented in such a heavily pretreated population of patients with ALL. The magnitude of OS benefit with brexucabtagene autoleucel in this patient population has not been achieved in previous trials in this setting, marking a significant advance for the field.

Acral, mucosal, and uveal melanomas are rare and deadly subtypes of melanoma. Treatment options are limited, and patients are often diagnosed at advanced stages. Scientists Dr. Cristina Puig-Saus and Dr. Daniel Shin of the UCLA Jonsson Comprehensive Cancer Center received a $1 million Translational Research Award from the U.S.

Department of Defense Melanoma Research Program to help advance the use of CAR T cells as a treatment for people with melanoma. Puig-Saus and Shin, whose labs are investigating how to improve the effectiveness of immunotherapies such as CAR T cell therapy for more people, will use the funds to establish a panel of acral, mucosal and uveal melanoma models, which to date are very limited, and will conduct preclinical studies to demonstrate the efficacy and improve the safety of tyrosinase-linked protein-1 (TYRP-1) CAR T cells in those models. TYRP-1 is highly expressed in 58.3% of acral melanomas, 60% of mucosal melanomas and 91.7% of uveal melanoma biopsies.

-- Trial will evaluate potential of next generation of influenza vaccines with combined mRNA technology and flu vaccine development expertise --

The FDA has approved lisocabtagene maraleucel (Breyanzi, Bristol Myers Squibb) as a new chimeric antigen receptor (CAR) T-cell therapy for adults with relapsed or refractory large B-cell lymphoma (R/R LBCL).

In September, the U.S. Food and Drug Administration suspended the Phase 1 trial evaluating Cellectis' investigational CAR T-cell therapy, UCARTCS1A, for the treatment of multiple myeloma for safety reasons after a patient who received the second lowest dose of the therapy died of cardiac arrest. UCARTCS1 cells are modified to carry an artificial receptor that targets the SLAMF7, or CS1, protein found at high levels on the surface of myeloma cells. The modified cells are developed in the laboratory and then infused into the patient to help fight the cancer. Cellectis has worked closely with the regulatory agency to meet its requirements and has modified the clinical protocol to increase patient safety. Recently, the FDA lifted the suspension of the Phase 1 trial for the UCARTCS1A CAR T cell therapy. The trial intends to recruit a total of 18 participants between the ages of 18 and 64 years at three sites in the United States: MD Anderson Cancer Center, Texas; Hackensack Meridian Health, New Jersey; and Weill Cornell Medicine, New York. To be eligible for the study, patients must have a confirmed diagnosis of active multiple myeloma and have relapsed after previous treatment.