Thread by @simocristea

New🔥TCR #CRISPR✂️ cancer #immunotherapy paper just out in @Nature by PACT Pharma & UCLA.

Why all the 📸headlines & press coverage?

This small proof-of-concept study can give us a glimpse into the future of cancer therapy.

Let’s unpack the details & relevance of this study👇🧵

First things first:

Here’s the link to the paper, which has been made public by @Nature in an unedited form (before official publication), due to its perceived immediate relevance to the research & clinical cancer communities.

nature.com/articles/s4158…

A. The context ✍️ of this work:

Most cancerous lesions have mutations in the 🧬of their cells.

Cancer cells (literally) display small pieces of cellular mutated proteins (called peptides) on their surface.

These peptides, called neoantigens, are important therapeutic targets.

How neoantigens are targeted:

1. major histocompatibility complex (MHC) molecules present a given neoantigen on the cell’s surface.

2. if any T cell can recognize the neoantigen via its receptor (TCR=T cell receptor), then it binds to it.

3. T cell destroys 🤺the mutated cell.

Many challenges to this process exist, including the fact that the binding TCR-MHC (or HLA, as human MHC=HLA) is very specific.

The human gene loci encoding the HLA are diverse, meaning that multiple different HLA phenotypes exist (each person has their own 🧬configuration 🧩).

Currently, most therapies are limited to patients with very few HLA phenotypes & are unavailable for the rest.

However: TCRs that bind to multiple HLA alleles could recognize neoantigens better & kill cancer cells more efficiently.

Hence: TCR engineering has huge potential🔮

Current TCR engineering approaches rely mostly on recombinanat viral vectors,which are hard to scale.

In turn, #CRISPR would facilitate simultaneous & more efficient knockouts of the endogenous TCR chains while inserting the transgenic TCR under the physiologic TCR promoter 🧬✂️

B. Solution💡proposed in this work:

An efficient method part of Adoptive Cell Therapy (immunotherapy) to:

1. isolate 🧫
2. engineer🧬✂️
3. insert back💉

multiple novel neoantigen-specific TCRs (neoTCRs) for 16 patients’ solid tumor mutations.

Patients treated dec2019-aug2022.

Let’s break down the steps of this process:

1. Identify expressed tumor mutations using bioinformatics pipelines 💻.

2. Prioritize neoantigen-peptide HLA candidate pairs per patient using bioinformatics & synthesize them (up to 104 per patient, corresponding to 34 unique HLAs).

3. Isolate & enrich CD8 Tcells from the patient’s blood.

4. Stain these T cells for the library created in step 2👆to isolate rare blood-circulating Tcells predicted to both recognize patient-specific antigens & bind to their HLA complexes (median/patient:8 TCRs for 5 antigens).

5. CRISPR engineer 🧬✂️ healthy donor CD8 & CD4 Tcells to mimic the identified rare cells by knocking out the endogenous TCR beta chain & inserting the transgenic TCR alpha & beta genes in the endogenous locus.

For more details on TCR structure,see this👇
nature.com/articles/s4157…

6. Test newly assembled neoTCR candidates for specificity of HLA binding (57% ok) & select confirmed neoTCRs for clinical manufacturing (max 3/patient).

Caution: verifying is a must,as previously found antigens can be lost (bc of high mutability, sublonality,loss of HLA allele).

7. Put the expression of the neoTCR constructs under endogenous promoter regulation & infuse 💉cells in patients (across patients, they make up from 2% to 47% of live cell product).

8. Characterize how the engineered neoTCR T cells work inside patients & assess efficacy.

C. Results 🏥: How do neoTCRs act in patients?

- they exist intra-tumorally, in close contact with cancer cells. Also present in metastases at higher freq. than baseline native T cells w same antigen-specific TCR

- their phenotype shifts from effectors to stem & central memory

However‼️

The given T cell dose was low ➡️ limited in-vivo expansion ➡️ low chance of clinical efficiency/benefit.

Disease outcome:
- only minor side effects
- 11/16 patients: disease progression
- 5/16 patients: stable disease at first tumor assessment (day 28 post-infusion)

Additional current hurdles 🏔️

1. Time⏳: the road to clinical implementation is very slow.

Screening ➡️ TCR discovery: median 167 days
+
Enrollment ➡️ dosing (product manufacturing): median 102 days

2. Cost💰: this therapy is very expensive per patient, hence cannot be scaled.

How to increase 📈 neoTCR efficacy?
- strengthen conditioning chemotherapy
- add IL-2
- increase neoTCR positive cells & decrease WT cells in final product

How to decrease 📉 time & cost?
- automation, e.g. establish existing libraries of common HLA alleles-neoantigens pairs.

D. Relevance🔮: why is this study important?

1. One of the most complex therapies ever attempted!

2. First time CRISPR-engineered TCRs are tested in human solid tumors

3. Probably a main therapy of the future & opens way to new clinical routes

4. Proof of biomedical progress

Takeaways ✍️

1. This small phase 1 study is about the feasibility & safety of non-viral precision genome engineering 🧬✂️ to generate personalized T cells with neoantigen-specific TCRs for Adoptive Cell Therapy.

Such an approach is much more efficient than virus-based vectors.

2. The therapy is safe, but also of limited clinical benefit, very lengthy, expensive & complex.

3. In the same time, it is an amazing 🤩 evidence of tens of years of research & clinical progress across many areas s.a. immunotherapy, sequencing, genome editing & bioinformatics.

4. There’s tremendous room for improvement! Now is the time to join forces & push forward to create version 2.0 of genetically engineered fighters🤺 to fight against a huge evil.

We don’t yet know how this fight ends. But a new road to walk this hard path has just been found 🌄