Tumor-infiltrating lymphocytes (TILs) have been shown to be effective against solid tumors in various clinical trials. Despite the enormous disease burden and large number of premature deaths caused by colorectal cancer (CRC), studies on TILs isolated from tumor tissue of CRC patients are still rare. To date, studies on ACT often lack controlled and comparable expansion processes as well as selected ACT-relevant T-cell populations.

We describe a procedure for generating patient-specific TILs, which are prerequisites for clinical trials of ACT in CRC. The manufacturing and characteristics of these TILs differ in important modalities from TILs commonly used for this therapeutic approach.

Tumor tissue samples were obtained from 12 patients undergoing surgery for primary CRC, predominantly with low microsatellite instability (pMMR-MSI-L). Tumors in the resected specimens were examined pathologically, and an approved volume of tumor tissue was transferred to a disposable perfusion bioreactor. Tissue samples were subjected to an automatically controlled and highly reproducible cultivation process in a GMP-conform, closed perfusion bioreactor system using starting medium containing IL-2 and IL-12. Outgrowth of TIL from tissue samples was initiated by short-term supplementation with a specific activation cocktail. During subsequent expansion, TILs were grown in IL2-enriched medium.

Expansion of TILs in a low-scaled, two-phase process in the Zellwerk ZRP bioreactor under hyperoxic conditions resulted in a number of approximately 2 ×109 cells. The expanded TILs consisted mainly (73%) of the ACT-relevant CD3+/CD8+ effector memory phenotype (CD45RO+/CCR7-). TILs harvested under these conditions exhibited high cytotoxic functional potential, which was confirmed upon nonspecific stimulation (IFN, TNF cytokine assay).

Verantwortliche:
Hendrik C. Albrecht, Jannis Schwanemann, Dirk Gustavus , Stephan Gretschel

Laufzeit:

07/2020 – 12/2022

Veröffentlichung:

In diesem Projekt werden die Resektate von Metastasen kolorektaler Karzinome (CRC) und Resektate von primären CRC als Ausgangs-Tumormaterial für die Isolation und Expansion tumor-infiltrierender Lymphozyten für eine Adoptive Zelltherapie (ACT) verwendet. Dabei werden verschiedene Methoden zur Optimierung der TIL-Kultivierung (Kombinationen von Aktivierungsfaktoren und Koaktivierungsfaktoren) untersucht.

Weiterhin werden die expandierten TIL-Populationen aus primären therapienaiven kolorektalen Karzinomen mit denen aus Metastasen kolorektaler Karzinome hinsichtlich Zell-Zahl, Phenotyp und Funktionszustand verglichen.

Weiterhin wird die spezifische Zytotoxizität der expandierten TIL durch Stimulation mit Zell-Suspensionen des jeweiligen Tumorausgangsmaterials analysiert.

Verantwortliche:
Hendrik C. Albrecht, Jannis Kröger, Evgueni Sinelnikov, Stephan Gretschel

Laufzeit:
11-2022 –

Förderung:
private Förderung Fa. KDH (6200EUR)