Immunotherapeutic Discovery & Development: Breakthroughs, Innovative Strategies, & Cell-based Approa

Author

Gaurav Agrawal, Ph.D., Head of Scientific Market Development at Eurofins DiscoverX®

EVOLUTION OF IMMUNOTHERAPY

While immunotherapy is a relatively new form of treatment, the evolution of immunotherapy spans centuries. It is marked by significant breakthroughs that have revolutionized our understanding of the immune system and its role in combating life-threatening diseases such as autoimmune disorders, allergies, and cancer.
 
The roots of immunotherapy can be traced back to the late 17th century when the concept of immunization emerged with the groundbreaking work of Dr. Edward Jenner (1) who developed the smallpox vaccine from cowpox lesions (see Figure 1.). While Jenner’s work predates the formal concept of immunotherapy as we know it today, his contributions to vaccination research had profound implications for the immunology field and the prevention of infectious diseases. Soon thereafter, another historic groundbreaking observation on tumor regression, post-surgical bacterial infections (2), laid the foundation for the concept that the immune system could be harnessed to fight cancer – establishing a branch of immunology now referred to as immuno-oncology (IO).
 
Figure 1. Significant breakthroughs in immunotherapy.
 
Figure 1. Significant breakthroughs in immunotherapy.
 
Significant strides in immunotherapy were made in the 20th century, particularly with the discovery of antibodies and their role in immune responses, setting the stage for targeted immunotherapy. Dr. Paul Ehrlich developed the concept of antibodies and called them the immune system’s “magic bullets” (3). Ehrlich’s work laid the groundwork for the understanding of humoral immunity, the branch of the immune system involving antibodies circulating in the blood. This concept became crucial in the development of different classes of antibodies, such as rituximab for certain cancers, showcasing the potential of harnessing the immune system to treat diseases (2).
 
The 21st century witnessed a significant leap in the field of cancer immunotherapy with the identification of immune checkpoint receptors and their crucial roles in cancer biology. This groundbreaking discovery was recognized with the Nobel Prize in Physiology or Medicine in 2018, awarded to Drs. James P. Allison and Tasuku Honjo (4). Their pioneering work elucidated how tumor cells ingeniously evade the immune system by exploiting some key checkpoint receptors. This led to the development of clinically approved antibody drugs like ipilimumab and nivolumab (Opdivo®) targeting checkpoint receptors CTLA-4 and PD-1, respectively. By effectively blocking their respective receptors, these antibodies unleash the immune system’s potent ability to recognize and eliminate cancer cells.
 
In more recent years, the immunotherapy landscape has expanded to include further innovative yet complex approaches such as cell and gene therapies (5). Chimeric antigen receptor T-cell (CAR-T) therapy has emerged as a revolutionary treatment. CAR-T involves genetically modifying a patient’s T cells to express a receptor that recognizes and targets cancer cells. This personalized approach has demonstrated success in certain blood cancers, such as leukemia and lymphoma, achieving durable responses and even leading some patients to complete remission. The approval of CAR-T therapies, such as KYMRIAH® and YESCARTA®, represents a paradigm shift in the treatment of certain hematologic malignancies.

INNATE AND ADAPTIVE IMMUNE SYSTEMS IN IMMUNOTHERAPY

Innate and adaptive immune systems play pivotal roles in the development of immunotherapeutic strategies to combat cancer and other life-threatening diseases (see Figure 2.). Innate immunotherapy focuses on augmenting the innate immune system’s inherent ability to identify and eradicate cancer cells. Cytokines like Interleukin-2 (IL-2) can directly activate innate immune cells, such as natural killer (NK) cells and macrophages, to enhance their anti-tumor activity (7).
Figure 2. Immune systems in immunotherapy
Figure 2. Immune systems in immunotherapy. Several established Immunotherapy approaches, including antibodies, checkpoint blockade, cytokines therapy, Fc effector function, vaccines, and cell and gene therapy exist.
 
Several other therapeutic strategies that harness innate immunity are actively being investigated. One approach involves targeting the axis between CD47, a cell-surface receptor on cancer cells that interacts with SIRPα, an inhibitory receptor on myeloid cells such as macrophages and dendritic cells (8). The SIRPα /CD47 interaction blocks the phagocytosis of cancer cells. By blocking this interaction, for example with an anti-CD47 antibody, these inhibitory signals are disrupted, allowing myeloid cells to recognize and eliminate cancer cells. Another strategy involves targeting ILT2/4, a pair of receptors on myeloid cells that also contribute to cancer immune evasion. Inhibiting ILT2/4 restores the ability of myeloid cells to identify and eliminate cancer cells. STING agonists, a class of innate immune activators, are promising therapeutics as they mimic the effects of natural ligands that bind to STING. The activation of the STING signaling pathway leads to the production of interferons and other immune-stimulating molecules, which can enhance the killing of cancer cells (9).
 
Conversely, adaptive immunotherapy aims to activate and expand adaptive immune cells to specifically target cancer cells. This can be accomplished through several methods, including checkpoint inhibitors, CAR-T cell therapy, bispecific antibodies, and antibody-drug conjugates (ADCs). In recent years, tumor vaccines have served as an excellent example of leveraging the adaptive immune system in immunotherapy. The tumor vaccines aim to activate and expand tumor-specific T cells and B cells, and they can use tumor-associated antigens, modified cancer cells, or dendritic cells pulsed with tumor antigens to induce an immune response against cancer (10).
 
While these two established immune systems present several therapeutic strategies independently, the synergistic application of innate and adaptive immunotherapy approaches is gaining momentum in cancer treatment, offering the potential for enhanced therapeutic outcomes and improved patient survival.

REGULATING T-CELL RESPONSES IS CENTRAL TO IMMUNOTHERAPY

Modulating T-cell responses is pivotal in immunotherapy where therapeutic development strategies often center around activating the immune system to identify and eliminate cancer cells. Blocking antibodies targeting co-inhibitory checkpoint receptors, such as pembrolizumab, are prominent examples of enhanced T-cell activity. Nevertheless, patients frequently fail to mount durable antitumor responses following checkpoint inhibitor (CI) immunotherapy due to transient and non-persistent T-cell activation. It is postulated that additional stimuli may be necessary to elicit a sustained immune response. An evolving approach involves the activation of co-stimulatory receptors to achieve a more enduring antitumor response, independently or in combination with CI therapy.
 
Agonistic antibodies are emerging as promising therapeutics for targeting co-stimulatory checkpoint receptors (see Figure 3.). Several clinical investigations are underway to evaluate the efficacy of agonistic antibodies targeting CD40, CD137, and CD28 to augment T-cell activation via co-stimulatory receptor engagement (11). In addition, there is also a growing focus on the potential of agonistic antibodies to selectively activate co-inhibitory checkpoint receptors as an avenue for suppressing inflammation in autoimmune diseases (12). Novel therapeutic strategies employing agonistic antibodies targeting immune checkpoint receptors, PD-1 and BTLA are currently undergoing clinical evaluation. However, their development often poses challenges as most in vitro assays fail to identify agonistic effects as they are unable to replicate the necessary physiologically relevant conditions.
Figure 3: Co-stimulatory and co-inhibitory receptors involved in T-cell activation
Figure 3: Co-stimulatory and co-inhibitory receptors involved in T-cell activation. Summary of numerous co-stimulatory and inhibitory receptors and their respective ligands that govern T-cell response and influence some of the key approved immunotherapeutic modalities.

HARNESSING CYTOKINES FOR IMMUNOTHERAPY

Targeting cytokines has emerged as a promising approach for cancer immunotherapy (13) (see Figure 4). IL-2, a naturally occurring cytokine, has been at the forefront of cytokine-based cancer therapies, activating T-cells to destroy tumor cells. Other similar cytokines, such as IL-15, enhance the anti-tumor activity of NK cells (14). IL-10, while not directly stimulatory, regulates immune responses, preventing excessive inflammation and suppressing anti-tumor immunity. Cytokines such as M-CSF and GM-CSF, that play essential roles in the development and function of macrophages and granulocytes, can be hijacked by tumor cells leading to the accumulation of tumor-promoting immune cells (15). Antibodies targeting CSF1R and GM-CSFR block their signaling pathways, thereby restoring the ability of macrophages and granulocytes to function against cancer. The potential of cytokines to harness the body’s innate immune defense system against cancer holds immense promise for the future of cancer treatment.
Figure 4. Cytokines involved in cancer immunotherapy
Figure 4. Cytokines involved in cancer immunotherapy. Cytokines like IL-2 and IL-15 promote the body’s immune cell development and proliferation to target and tumor cells. However, other cytokines, like GM-CSF and M-CSF, play an opposing role by modulating tumor-associated macrophages (TAM) and promote tumor cell proliferation in the tumor microenvironment. Antibodies blocking such cytokines present a promising avenue to prevent tumor growth.
 
In autoimmune disorders, an imbalance in cytokine production is often the underlying cause of the disease. For instance, in rheumatoid arthritis, excessive production of TNF-α contributes to joint inflammation and destruction. Targeting specific cytokines through immunotherapy effectively modulates the immune response and alleviates the symptoms of autoimmune disorders. For example, several anti-cytokine therapeutics are clinically approved for psoriatic arthritis and Crohn’s disease (16).
 
While cytokine-based immunotherapy has demonstrated efficacy in treating various autoimmune disorders, challenges remain. One concern is the potential for side effects, such as increased susceptibility to infections. Additionally, not all patients with autoimmune diseases respond favorably to cytokine-based therapy.
 
As research ventures deeper into the intricate workings of the immune system and its complex interplay with cancer, we can anticipate even more sophisticated and targeted immunotherapy strategies, propelling us closer to the ultimate triumph over cancer. As we delve further into the 21st century, ongoing research continues to uncover new possibilities for immunotherapy, including the exploration of novel targets, the development of combination therapies, and the application of these approaches to a broader spectrum of cancers. The evolution of immunotherapy represents a triumph of scientific discovery and has ushered in a new era of cancer treatment, offering hope and tangible results for patients who were once faced with limited options.

CELL-BASED ASSAYS PLAY AN ESSENTIAL ROLE IN IMMUNOTHERAPY DRUG DEVELOPMENT

The development of immunotherapy modulators necessitates the implementation of phase-specific assay solutions. During the initial stages, a vast array of molecules is screened using rapid methods in a high-throughput format. Biochemical assays, owing to their relative simplicity, are often employed in this early phase. However, drug candidates can exert their effects on living organisms through a multitude of mechanisms that biochemical assays fail to capture. These limitations have spurred an increased reliance on biologically relevant cell-based assays that provide a more representative assessment of an organism’s response to a drug. Cell-based assays can also be miniaturized and adapted to a high-throughput format for primary screening programs.
 
For biotherapeutic drugs such as monoclonal antibodies, a mechanism of action (MOA)-reflective, physiologically relevant assay is crucial for accurately determining the potency and stability of the drug product. Consequently, as the drug progresses through the various phases of clinical development, the criteria for selecting a cell-based assay becomes increasingly stringent. There is a growing trend of utilizing cell-based assays in immunogenicity studies (as part of the clinical trial) to ascertain whether anti-drug antibodies produced in the treatment-receiving subjects neutralize the drug. Thus, the development of cell-based assays for characterization, potency testing, and neutralizing antibody detection is critical to meet regulatory expectations and is widely recognized as a challenging and time-consuming process due to its inherent complexities. If the development of cell-based assays is deferred to later stages of clinical development (as often noted by regulatory agencies), the commercial release of the drug to the market may be delayed owing to a lack of supporting characterization or potency testing data.
 
To circumvent these development challenges and accelerate drug development programs, implementing commercially available pre-qualified assay platforms can significantly conserve time and resources. Eurofins DiscoverX® addresses this need by offering the industry’s most comprehensive portfolio of optimized, MOA-reflective cell-based assays for immune checkpoint modulators, cytokines, and other targets, available in phase-appropriate formats.

Select Related Resources

 

REFERENCES

  1. Jenner, E. An inquiry into the causes and effects of the variolae vaccinae: a disease discovered in some of the western counties of England, particularly Gloucestershire, and known by the name of the cow pox. Medicine in the Americas, 1610-1920 (1798).
  2. Starnes, C.O. Coley’s toxins in perspective. Nature. 1992 May 7; 357 (6373): 11-2.
  3. Ehrlich, P. Experimental Researches on Specific Therapy, The Collected Papers of Paul Ehrlich, Elsevier, 106–117 (1960).
  4. Guo, Z.S. The 2018 Nobel Prize in medicine goes to cancer immunotherapy (editorial for BMC cancer). BMC Cancer. 2018 Nov. 12; 18 (1): 1086.
  5. Porter, D.L, Hwang, W.T., Frey, N.V., Lacey, S.F., Shaw, P.A., et al., Chimeric antigen receptor T cells persist and induce sustained remissions in relapsed refractory chronic lymphocytic leukemia. Sci Transl Med. 2015 Sept. 2; 7 (303): 303ra139.
  6. Medzhitov, R. Toll-like receptors and innate immunity. Nat Rev Immunol 1, 135-145 (2001).
  7. Striz, I., Brabcova, E., Kolesar, L., Sekerkova, A. Cytokine networking of innate immunity cells: a potential target of therapy. Clin Sci (Lond). 2014 May; 126 (9): 593-612.
  8. Qu, T., Li, B., Wang, Y. Targeting CD47/SIRPα as a therapeutic strategy, where we are and where we are headed. Biomark Res 10, 20 (2022).
  9. Zhu, Y., An, X., Zhang, X. et al. STING: a master regulator in the cancer-immunity cycle. Mol Cancer 18, 152 (2019).
  10. Le, I., Dhandayuthapani, S., Chacon, J., Eiring, A.M., Gadad, S.S. Harnessing the Immune System with Cancer Vaccines: From Prevention to Therapeutics. Vaccines (Basel). 2022 May 21; 10 (5): 816.
  11. Mayes, P., Hance, K., Hoos, A. The promise and challenges of immune agonist antibody development in cancer. Nat Rev Drug Discov 17, 509–527 (2018).
  12. Grebinoski, S., Vignali, D.A. Inhibitory receptor agonists: the future of autoimmune disease therapeutics? Curr Opin Immunol. 2020 Dec.; 67: 1-9.
  13. Berraondo, P., Sanmamed, M.F., Ochoa, M.C., et al. Cytokines in clinical cancer immunotherapy. Br J Cancer 120, 6-15 (2019).
  14. Yang, Y., Lundqvist, A. Immunomodulatory Effects of IL-2 and IL-15; Implications for Cancer Immunotherapy. Cancers (Basel). 2020 Nov. 30; 12(12): 3586.
  15. Ushach, I., Zlotnik, A. Biological role of granulocyte macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF) on cells of the myeloid lineage. J Leukoc Biol. 2016 Sept.; 100 (3): 481-9.
  16. Uricoli, B., Birnbaum, L.A., Do, P., Kelvin, J.M., Jain, J., et al.. Engineered Cytokines for Cancer and Autoimmune Disease Immunotherapy. Adv Healthc Mater. 2021 Aug.; 10 (15): e2002214.
  17. United States Pharmacopeia (2023). General Chapter, (1032) Design and Development of Biological Assays. USP-NF. Rockville, MD: United States Pharmacopeia.