Next-Generation Sequencing-Guided Cancer Immunotherapy 

Diagnomics works exclusively with Multimeric Biotherapeutics to use advanced bioinformatics with Next-Generation Sequencing (NGS) and UltraCD40L™ to enable the Adoptive Cell Therapy (ACT) of cancer.

 

History
Melanomas and many other tumor types contain Tumor Infiltrating Lymphocytes (TILs) that recognize tumor antigens. TILs can be expanded ex vivo by culturing them in presence of interleukin 2 (IL2). When these TILs are re-infused back into the patient along with IL2 treatment, about 25% of patients are cured. However, treatment mortality is about 10-15% and most treated patients develop serious form of autoimmune disease.

Fig.1: Schematic of TIL therapy

 

Adoptive Cell Therapy (ACT)
Infusions of Cytotoxic T Lymphocytes (CTLs) can eliminate tumors in humans. Fig. 2 shows the eradication of melanoma metastases to the liver following an infusion of CTLs cultured from tumor-infiltrating lymphocytes (TILs).

 

Newer forms of ACT
Ex vivo culturing of immune cells with tumor antigen (example: Dendreon). Ex vivo culturing of anti-tumor T cells using dendritic cells as antigen-presenting cells (APCs) (examples: Pacific Northwest for glioblastomas and Geron for melanomas). In addition, ex vivo culture of anti-tumor T cells using Epstein-Barr Virus (EBV)-transformed B cells as APCs is being developed (example: Cell Medica). The common goal is to find faster, better and cheaper ways of presenting tumor antigens to T cells.

 

Our approach: Multimerized TNF Super Family Receptors (TNFSRs)
Receptor clustering is needed to activate TNFSF receptors. For best efficiency a many-trimer multimer (>2 trimers) is required.

 

CD40 Ligand (CD40L)
CD40 Ligand is the strongest immune stimulating molecule made by the human body

 

Creation of UltraCD40L™
UltraCD40L™ is a fusion protein constructed by joining the body of SP-D to the extracellular domain of CD40L.

 

CD40L-stimulated B cells
CD40L-stimulated B cells can be used to generate anti-tumor CD8+ cytotoxic T cells (CTLs)

 

Growth and activation of primary B cells
Primary B cells harvested from patient's own blood grow indefinitely when supplied with CD40L + IL4

Methods: 50 ml of blood are obtained from cancer patients and cultured with 3T3 cells expressing membrane CD40L plus IL-4. By re-stimulating every 5 days, unlimited numbers of B cells could be generated in vitro.

von Bergwelt-Baildon, M. et al. (Dana-Farber group) Blood 99:3319-3325, 2002.

 

UltraCD40L™ and Adoptive Immunotherapy
UltraCD40L™ can be used to grow B cells for adoptive immunotherapy.

Growth of primary human B cells using UltraCD40L™ + IL4

Comparison of B cells grown with UltraCD40L™ versus Dendreon Provenge™

NGS Reveals Many Tumor-Specific Antigens for Cancer Immunotherapy

Cancer cells contain numerous mutations, many of which encode mutated peptides that be used for a specific immune attack on the tumor cells while sparing normal host cells. We are using specifically developed computer algorithms to analyse the epitope landscape of many different cancer types isolated from primary tumors and metastases.

 

Overall Conclusions


Abbreviations used

DCs - Dendritic Cells
APCs - Antigen Presenting Cells
IL2 - Interleukin 2
IL4 - Interleukin 4
NGS - Next Generation Sequencing
CTLs - CD8+ cytotoxic T cells
TNFSF - Tumor Necrosis Fractor Super Family