Overview

On November 2, commercial insurance catalog negotiations officially began, with five CAR-T therapies entering the negotiation list: Fosun Kite’s Axicabtagene Ciloleucel, JW Therapeutics’ Relmacabtagene Autoleucel, IASO Bio’s Equecabtagene Autoleucel, CARsgen Therapeutics’ Zevo-Cabtagene Autoleucel, and Abogen’s Nacacabtagene Autoleucel.

As a supplement to the basic medical insurance system, the innovative drug catalog under commercial insurance adopts a negotiated pricing mechanism instead of deep price cuts, reserving space for the development of innovative therapies.

Since the beginning of this year, the biopharmaceutical industry has witnessed an intense “arms race.” AstraZeneca, AbbVie, Gilead, and BMS have each spent hundreds of millions of dollars to acquire early-stage in vivo CAR-T companies. In sharp contrast, traditional autologous and allogeneic (off-the-shelf) CAR-T therapies are showing growth bottlenecks — complex manufacturing, high cost, and limited efficacy against solid tumors.

Schematic diagram of CAR lentiviral vector structure

Reconstructing the Treatment Logic: Precision Delivery and Cellular Engineering

Although traditional ex vivo CAR-T therapy shows great potential, it has always been limited by three major challenges:

Complex process: 

It requires blood collection, ex vivo T-cell modification and expansion, which are time-consuming and rely on specialized facilities.

Limited manufacturing capacity: 

The personalized nature of production makes large-scale manufacturing difficult, restricting patient accessibility.

Preconditioning dependency: 

Patients need to undergo chemotherapy to deplete immune cells before infusion, increasing infection risks.

In contrast, in vivo CAR-T delivers CAR genes directly into the body via specific carriers, precisely targeting T cells, NK cells, or even hematopoietic stem cells for in situ genetic modification. This strategy retains the precision killing power of CAR molecules while achieving scalability — breaking through the long-standing limitations of traditional CAR-T therapies.

Technical Routes:

“Long-acting” Lentiviral Vector VS “Controllable” LNP Vector

Lentiviral vectors – One-time treatment for long-term protection:

Lentiviral vectors integrate the CAR gene stably into the T-cell genome, enabling durable CAR expression and prolonged tumor-killing capability. The core technologies lie in T-cell–specific targeting, transduction efficiency, and safety regulation.

01. T-cell targeting transformation: from broad tropism to T-cell specificity

Traditional lentiviruses use broadly tropic envelope proteins (such as VSV-G), which can infect hepatocytes and endothelial cells. Dual engineering — detargeting + retargeting — achieves T-cell–specific delivery:

Envelope protein engineering: 

Replace VSV-G with a CD28L chimeric envelope, fusing the T-cell co-stimulatory ligand CD28L. This specifically binds to the CD28 receptor on T cells, increasing transduction efficiency 30-fold over wild-type lentivirus while reducing off-target hepatocyte infection to <0.1%.

Genomic element optimization: 

Introduce an RACR/CAR system where the lentiviral vector carries both a “reverse responsive element” and the CAR gene. Without drug administration, CAR expression remains silent and T cells are quiescent. After oral administration of the small molecule AP21967, the responsive element changes conformation to activate CAR expression, peaking in 48 hours and silencing again within 7 days after drug withdrawal — reducing CRS incidence by 60%.

02. Transduction efficiency optimization: high-efficiency T-cell modification at low doses

Lentiviral transduction efficiency depends on viral titer, T-cell activation, and co-stimulatory signals. The following strategies enable breakthroughs:

High-titer production: 

By adopting a “suspension + serum-free culture” system and optimizing HEK293T transient transfection, viral titers increased from 1×10⁸ TU/mL to 5×10⁹ TU/mL. This supports effective transduction at one-tenth the standard dose, minimizing off-target risks.

T-cell activation synergy: 

Embedding an IL-2 mimetic peptide within the vector activates T-cell proliferation signals, increasing the proportion of G1-phase T cells from 35% to 62%, boosting transduction efficiency 1.8×. Meanwhile, integrating 4-1BBL into the envelope provides a second co-stimulatory signal by binding 4-1BB on T cells, enhancing survival and persistence — with CAR-T cells persisting up to 12 months in cynomolgus monkey models.

03. Clinical safety regulation: enabling long-lasting T-cell killing

Hematologic malignancies — Sustained response at low dose: 

With CD45-ligand lentiviral delivery of CD19 CAR at 1×10⁸ TU/kg, peripheral T-cell transduction reached 63%, tumor burden dropped by 92% in lymphoma mouse models, and IL-6 levels were 50% lower than ex vivo CAR-T.

Solid tumors — Enhanced tumor penetration: 

Lentiviral vectors co-delivering “BCMA-CAR + TGFβ-neutralizing peptide” in multiple myeloma models neutralized TGFβ-mediated immunosuppression, enabling CAR-T infiltration deep into tumor cores. The Treg proportion in tumor microenvironment decreased from 32% to 11%, improving killing efficiency by 2.5×.

LNP Vector – “Safe, Controllable, and Flexible”

CAR mRNA does not integrate into the genome, naturally avoiding genotoxic risks. mRNA-LNP uses “antibody-targeted LNPs” to deliver CAR mRNA directly to T cells in vivo, achieving in situ T-cell reprogramming. The core technologies involve Ab-LNP targeting, formulation, and clinical validation.

01. Ab-LNP specific targeting: precise recognition of T cells

The key to Ab-LNP lies in antibody modification that forms a specific bridge between LNPs and T-cell receptors. The process includes: antibody conjugation of CD3/CD8-specific antibodies to LNP surfaces → specific binding to T-cell receptors → endocytosis and mRNA release → high-efficiency CAR expression within cytoplasm.

02. Ab-LNP formulation: enhancing T-cell transduction efficiency

Ab-LNP preparation optimizes antibody density, ionizable lipid pKa, particle size, and mRNA loading to maximize transduction efficiency.

03. Ab-LNP clinical validation: safety and efficacy

mRNA-LNP–based in vivo CAR-T technology has achieved multiple breakthroughs:

• Over 90% clearance of hematologic malignancies without hepatic toxicity;
• Significant symptom relief in autoimmune diseases (e.g., lupus) with no neurotoxicity;
• Effective solid tumor control by incorporating chemokine mRNA to improve infiltration — combined with PD-1 inhibitors, tumor suppression reached 89%.

Risks and Outlook: The “Last Mile” from Lab to Bedside

Although in vivo CAR-T holds great promise, several barriers remain before clinical translation:

Lentiviral long-term risks: 

Persistent CAR expression may cause chronic B-cell depletion; genomic integration risks require long-term monitoring.

mRNA-LNP short-lived expression: 

Transient CAR expression limits long-term tumor control, requiring repeat dosing.

Cell selectivity: 

Off-target transduction of non-intended cells can reduce efficacy or cause organ toxicity.

The industry is working to overcome these issues: lentiviral vectors are being optimized with improved envelope design to reduce immunogenicity and integration risks, while mRNA-LNP systems are leveraging circular mRNA to extend expression duration. Both are evolving toward more precise targeting and longer-lasting therapeutic effects.

In vivo CAR-T therapy marks a key evolutionary step in cell therapy — shifting from personalized, case-by-case treatment to standardized and scalable therapeutic models.
This technological revolution not only addresses the cost and accessibility challenges of traditional CAR-T therapy but also heralds a new era in cell-based immunotherapy.

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Bibliography

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