Genetic Engineering Publications - GEG Tech top picks

Glioblastoma (GBM) is the most common and aggressive type of cancerous brain tumor in adults. People with GBM generally expect to live 12 to 18 months after diagnosis. Despite decades of research, there is no known cure for GBM, and treatments have only a limited effect on extending an individual's life expectancy. However, researchers have tested a technology that delivers CAR-T cells targeting two proteins commonly found in brain tumors: epidermal growth factor receptor (EGFR), estimated to be present in 60% of all GBMs, and interleukin-13 receptor alpha 2 (IL13Rα2), which is expressed in over 75% of GBMs. While CAR-T cell therapy for blood cancers is usually administered intravenously, the researchers administered these dual-targeted CAR-T cells intrathecally, by injection into the cerebrospinal fluid, so that the modified cells could reach the tumors more directly in the brain. Magnetic Resonance Imaging scans taken 24 to 48 hours after administration of dual-targeted CAR-T cells targeting EGFR and IL13Rα2 revealed a reduction in tumor size in all six patients, and these reductions were maintained up to several months later in a subgroup of patients. 

Genetic control in mammalian cells is essential for the development of safe and effective gene therapies. Current methods are associated with certain drawbacks, such as undesirable immunological responses, limited efficacy and overexpression of therapeutic genes. Current gene transfer technologies, such as adeno-associated viruses (AAVs), have difficulty achieving conditional and reversible gene control. Toxic ligands, leakage and high ligand concentrations, as well as small dynamic range are some of the limitations associated with current RNA-based systems. In a recent study researchers describe the pA regulatory system, inserted into cells by CRISPR-Cas9, based on a ribonucleic acid (RNA)-based switch to regulate mammalian gene expression by modulating the cleavage of a synthetic polyA signal (PAS) at a transgenic 5' untranslated region. (UTR). This technique differs from traditional riboswitch systems in that the PAS is present in the 5' UTR, combines the effects of numerous aptamers and uses two processes of Tc binding and alternative splicing. However, the new system can only use Tc as an inducing ligand, which cannot effectively penetrate all body tissues.

The race to the clinic is reviving for a ready-made alternative to autologous CAR-T cell therapy, even as concerns about chromosomal abnormalities persist. The Advanced Regenerative Medicine Therapy designation, which makes the therapy eligible for accelerated approval, will also help remove a veil that has hung over standard CAR-T cell therapies since October, when the FDA put all trials of competitor Allogene Therapeutics on hold following the detection of a chromosomal abnormality in a patient who received ALLO-501A in a Phase 2 trial. The FDA's green light for CRISPR Therapeutics dispels broader concerns that the agency views this type of genotoxic safety event as an intractable problem for the entire class of allogeneic CAR-T therapies. Today, many companies are eliminating loci associated with the MHC-I to avoid host T cell recognition of transplanted CAR-T cells. Companies also equip their T cells with a variety of safety switches and performance enhancers.

However, as the complexity of the assembly increases, the risk of off-target effects also increases. This may be important from a safety perspective, given that most cancers lack unique antigens. Achieving rapid remission and re-dosing if necessary, can minimize the toxic effects that CAR-T cells can have on healthy tissues expressing the targeted antigen.

According to Jing Pan's article in the Journal of Clinical Oncology titled: Donor-derived CD7 chimeric antigen receptor T cells for T-cell acute lymphoblastic leukemia: first-in-human phase I trial, CAR T cells are reported to be remarkably effective in patients with B-cell acute lymphoblastic leukemia (ALL) but have not been successful to date in patients with T-cell ALL (T-ALL). Now, data from Pan and colleagues demonstrate the safety and impressive short-term efficacy of allogeneic donor-derived anti-CD7 CAR T cells in an early phase clinical trial involving patients with relapsed and/or refractory T-ALL.

In a paper now published in the journal Cell Stem Cell, the authors describe how the cells showed enhanced "anti-tumor activity" in mice with ovarian cancer seeded from human cancer cells.

A chimeric antigen receptor system that can integrate signals from multiple antigens
and fine-tune T cell activation in a cell-type-specific manner holds promise for enhancing
the safety and specificity of CAR T cell therapies for cancer treatment.

Here, the authors develop programmable, sequence-specific antimicrobials using the RNA-guided nuclease Cas9 delivered by a bacteriophage. They show that Cas9, reprogrammed to target virulence genes, kills virulent, but not avirulent, Staphylococcus aureus. This technology creates opportunities to manipulate complex bacterial populations in a sequence-specific manner.

In the last decade, new technologies such as optogenetics, chemogenetics and the CRISPR-Cas system have begun to transform how biologists understand the finer details associated with sleep-wake regulation. Here, the authors detail how some of the newest technologies are being applied to understand the neural circuits underlying sleep and wake.

In this chapter, the authors discuss the current state of epigenomic profiling and how functional information can be indirectly inferred. They also present new approaches that promise definitive functional answers, which are collectively referred to as 'epigenome editing'. In particular, they explore CRISPR-based technologies for single-locus and multi-locus manipulation. Finally, they discuss which level of function can be achieved with each approach and introduce emerging strategies for high-throughput progression from profiles to function.

A modified version of Cas9 with a fusion of the hormone-binding domain of the estrogen receptor allows reversible control of Cas9 activity with high efficiency at multiple loci with 4-hydroxytamoxifen treatment.

In this work, the authors established neuronal models of FTLD-Tau by Neurogenin2-induced direct neuronal differentiation from FTLD-Tau patient iPSCs. They found that FTLD-Tau neurons, either with an intronic MAPT mutation or with an exonic mutation, developed accumulation and extracellular release of misfolded tau followed by neuronal death, which we confirmed by correction of the intronic mutation with CRISPR/Cas9. FTLD-Tau neurons showed dysregulation of the augmentation of Ca2+ transients evoked by electrical stimulation.

This FTLD-Tau model provides mechanistic insights into tauopathy pathogenesis and potential avenues for treatments.

Axi-cel CAR T therapy targets the CD19 molecule on large B-cell lymphoma cells. The ZUMA-7 trial demonstrated that axi-cel reduced the risk of disease progression, need for retreatment or death by 60% compared with standard therapy. Despite these positive results in terms of event-free survival and overall survival, some patients did not respond well to treatment or relapsed rapidly after treatment. Researchers analyzed tumor gene expression patterns from patient samples and determined that a B-cell gene expression signature and CD19 protein expression were significantly associated with improved event-free survival for patients treated with axi-cel but not with standard therapy. Patients with lower tumor cell levels of CD19 showed gene expression patterns associated with immunosuppression. These observations suggest that the tumor immune environment may play an important role in regulating axi-cel treatment and outcome. Furthermore, biomarkers associated with improved axi-cel treatment outcomes decreased as patients received more treatments, suggesting that receiving axi-cel as part of earlier treatment lines is essential to ensure better patient outcomes.

Immunotherapy, particularly CAR T-Cell cancer therapy, extends the lives of many patients. But sometimes the therapy randomly migrates to places it shouldn't go, sneaking into the lungs or other non-cancerous tissue and causing toxic side effects. However, a team of researchers has discovered the molecule responsible for guiding T cells to tumors, setting the stage for scientists to improve the revolutionary treatment. Their discovery of the crucial migration control gene that expresses ST3GAL1 is the result of "unbiased genomic screening": researchers used a state-of-the-art CRISPR technique to edit thousands of genes expressed in T cells, then tested the migration control capabilities of these genes, one by one over a period of nearly four years, in mouse models. The next step is to find a drug that can manipulate the key T cell protein, ST3GAL1. If the study progresses as planned, such a drug could be added to the CAR T-cell regimen to ensure that the T cells reach their targets

In the new study, Dr. Sykulev and colleagues in Takami Sato's lab engineered CAR-T cells to recognize an antigen on melanoma cells called high molecular weight melanoma-associated antigen (HMW-MAA). Melanoma cells express varying amounts of HMW-MAA on their cell surfaces. In their research, the researchers evaluated the extent to which CAR-T cells killed melanoma cells. They found that CAR-T cells effectively killed melanoma cells expressing high levels of HMW-MAA, but not those with lower levels of the antigen. The researchers then tested how well another type of immunotherapy, known as TCR-T cells that uses T cells engineered to express a specific T-cell receptor, killed the target cells. When the researchers treated melanoma cells with TCR-T cells, they found that the treatment readily killed tumor cells, even in melanoma cell lines that expressed far less HMW-MAA antigens than needed for CAR-T. The comparison thus revealed that TCR-T is superior to CAR-T therapy for melanoma.

The "T-Cell Immunotherapy Market, 2018-2030 (3rd edition)" report features an extensive study of the current market landscape and the future potential of T-cell immunotherapies (focusing particularly on CAR-T therapies, TCR therapies and TIL therapies). One of the key objectives of the study was to review and quantify the future opportunities associated with the ongoing development programs of both small and big pharmaceutical firms.

The methods present in this article decouple aspects of kinetic and thermodynamic properties of the Cas9–DNA interaction and broaden the toolkit for investigating off-target binding behavior.

Using CRISPR/Cas9 technology, researchers have devised a method to deliver a CAR gene to a specific locus, TRAC, in T cells. This targeted approach yielded therapeutic cells that were more potent even at low doses; in a mouse model of acute lymphoblastic leukemia, they outperformed CAR T cells created with a randomly integrating retroviral vector.

Here, the scientists engineer the Light Plate Apparatus (LPA), a device that can deliver two independent 310 to 1550 nm light signals to each well of a 24-well plate with intensity control over three orders of magnitude and millisecond resolution. Signals are programmed using an intuitive web tool named Iris. All components can be purchased for under $400 and the device can be assembled and calibrated by a non-expert in one day.