Essay: Immunotherapy

Each cancer and tumor type is unique as is the individual in which it develops. Some cancers do not respond to established cancer treatments, such as chemotherapy or radiation therapy, because tumor cells have developed ways to evade the immune system by taking advantage of peripheral tolerance. Recent advances in immunotherapy have given rise to drastic advances in cancer treatment that allow targeted, tumor-cell-specific effects. These advances include immunomodulation checkpoint PD-1/PD-L1 and CTLA-4 inhibitors along with tumor associated antigen specific CAR-T-cell based therapies.
PD-1/PD-L1 based therapies use inhibitors that prevent the association of programmed death-ligand 1 (PD-L1) with its receptor programmed cell death protein 1 (PD-1) which are proteins present on the surface of specific cells. PD-1 is an immune checkpoint that prevents unwarranted immune responses to antigens and protects against autoimmunity. An immune checkpoint is a molecule on immune cells that needs to be activated to start an immune response. This allows for the body to discriminate between regular cells in the body (“self”) and foreign entities. When PD-1 expressed on T-cells encounters its ligands, its functional activities are reduced, including proliferation, cytokine secretion and cytolytic activity. This suppression of the immune system limits the killing of bystander host cells and prevents autoimmune disease.
PD-1 is expressed on a variety of immune cells including monocytes, T and B cells, dendritic cells and tumor infiltrating lymphocytes. PD-1 has two ligands: PD-L1 and PD-L2. PD-L1 is typically expressed on antigen presenting cells (APCs) and tumor cells. The binding of PD-L1 to PD-1 initiates antigen receptor signalling transduction leading to the destruction of T-cell metabolism and ultimately eliminates the immune function of T-cells. Some cancer cells have large amounts of PD-L1 which helps them evade immune attack. PD-L2 on the other hand is expressed on dendritic cells and monocytes and binds with a higher affinity to PD-1 than PD-L1.
Loss of immunologic control is a hallmark of cancer and the engagement of PD-L1 with PD-1 prevents the immune system from attacking tumor cells. Monoclonal antibodies (mAbs) that target either PD-1 or PD-L1 block this interaction and boost the body’s natural immune response against cancer cells. These antibodies act as tumor suppressing factors and enhance the antitumor immune response. This immune response starts with tumor cells producing mutated antigens that dendritic cells use to prime T-cells and stimulate the activation of cytotoxic T-cells. Activated T-cells then travel to and infiltrate the tumor environment and bind to the cancer cells. The bound effector T-cells then release cytotoxins, which induce apoptosis in their target cancer cells.
Typically, antibodies directed towards PD-1 are not preferred as they prevent its binding to both ligands which results in higher toxicity. PD-L1 inhibition as opposed to PD-1 inhibition blocks only PD-1:PD-L1 interactions while preserving PD-1:PD-L2 interactions. This provides more target specific signaling. PD-L1 also binds to CD80 to deliver inhibitory signals to T-cells so its inhibition prevents reverse signaling and resulting T-cell downregulation.
MAbs can significantly reduce toxicity while shrinking solid tumors, suppressing metastasis, and improving overall patient survival. Current FDA approved PD-1/PD-L1 inhibitors include Nivolumab and Pembrolizumab which are PD-1 inhibitors and Atezolizumab which is a PD-L1 inhibitor. These are currently the frontline treatments for metastatic melanoma, non-small cell lung cancer, renal cell carcinoma and urothelial cancer.
Immune mediated adverse reactions describe the side effects of immunotherapy. PD-1/PD-L1 inhibitors can have adverse reactions that lead to immune system dysregulation which can cause autoimmune disease-like symptoms. Classic chemotherapy toxicities (fatigue, anorexia, nausea, diarrhea), are also seen in patients treated with PD-1/PD-L1 inhibitors. These adverse effects are likely due to off-target interactions.
PD-1/PD-L1 inhibitors are closely related to CTLA-4 inhibitors as both PD-1 and cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) are expressed on activated T cells. The difference between the two inhibitors is the phase of the immune response in which they are active. While PD-1 inhibitors target T-cell activation in the tumor environment, anti-CTLA-4 antibodies centrally affect T-cell priming.
CTLA-4 is an immune checkpoint that stops and inactivates autoreactive T-cells that are unable to differentiate self from non-self, at the initial stage of activation in the lymph nodes. Activation of the CTLA-4 receptor downregulates any immunogenic response by binding to its ligands CD80 and CD86. The CTLA-4 receptor is expressed by both CD4+ (helper) and CD8+ (cytotoxic) T-cells while its ligands are present on the surface of APCs. CTLA‐4 is also expressed on regulatory T cells which suppress the immune response in the tumor microenvironment. Due to a variety of targets, a CTLA‐4 blockade results in a broad, nonspecific activation of an immune response.
CTLA-4 inhibitors are mAbs that attach and block CTLA-4 from binding to its ligands, which boosts the immune response against cancer cells. Blocking CTLA-4 induces an antitumor immune response by promoting the activation and proliferation of tumor specific T-cells. This blockade is also thought to promote the generation of memory T-cells which provides a long term antitumor response.
The mechanism of action behind this effect is explained by the fact that CTLA-4 is a CD28 homolog that binds with higher affinity to CD80 (B7.1) and CD86 (B7.2). The relative amount of CD28:B7 binding versus CTLA-4:B7 binding determines whether a T-cell will undergo activation. When CTLA-4 and its ligands bind, no stimulatory signal is produced. This competitive binding can prevent the stimulatory signal normally provided by CD28/B7 binding. Some evidence suggests that CTLA-4 binding to B7 may also produce inhibitory signals that counteracts the stimulatory signals from CD28:B7 binding.
Ipilimumab is an FDA approved CTLA-4 inhibitor which is a selective human IgG1 mAB used for the treatment of melanoma. It is currently undergoing clinical trials for the treatment of non-small cell lung carcinoma, small cell lung cancer, bladder cancer and metastatic prostate cancer. Like all immune checkpoint inhibitors, CTLA-4 inhibitors are associated with immune mediated toxicities, most of which can be managed successfully with corticosteroids. Commonly reported immune mediated adverse reactions include rash, itching and gastrointestinal disorders.
The CTLA-4 and PD-1/PD-L1 pathways have a similar negative effect on T-cell activity however the timing, signaling mechanism and location of immune inhibition by these two checkpoints differs. CTLA-4 is confined to T-cells whereas PD-1 is expressed on activated T cells, B cells and myeloid cells. While CTLA-4 functions during the priming phase of T-cell activation in lymph nodes, PD-1 functions during the effector phase largely in peripheral tissue. Recent studies show an enhanced antitumor response using a dual blockade compared with single agent blockade. This indicates that a blockade of both the CTLA-4 and PD-1/PD-L1 pathway could restore the immune responses of previously activated T-cells and reduce regulatory T-cell-mediated immunosuppression.
CAR-T-cell therapy does not use immune checkpoints as an alternative cancer treatment, but rather redesigns and redirects a patient’s own T-cells to specifically target and destroy tumor cells. Tumors use a variety of mechanisms to neutralize immune attacks. They can downregulate MHC molecule expression or disrupt antigen processing and presentation machineries. Chimeric antigen receptor (CAR) T-cell therapy use genetically engineered T-cells to find and destroy tumor cells in the body.
CAR-T cells are engineered to express synthetic receptors that redirect T-cells to surface antigens for tumor elimination. A CAR is a fusion protein composed of 3 domains. These include an extracellular single chain variable fragment that serves as the target binding domain, a transmembrane spacer domain and an intracellular signaling domain with cytoplasmic proteins that provide T-cell activation signals. These domains allow CAR-T cells to recognize cell surface tumor antigens leading to antigen-specific T-cell activation, proliferation, and cytokine production. Many CARs are also designed with elements that enhance T-cell persistence and activity.
After a CAR is designed, it is introduced into T-cells. Initially, through leukapheresis, peripheral blood mononuclear cells from a patient are collected and the blood is separated into different components. Once the immune cells are sequestered, the CAR constructs are transfected into T-cells using plasmid transfection, mRNA or viral vector transduction. More recent approaches to genetically altered T-cells include gene editing tools like CRISPR that create a double stranded break at a particular site within the genome and introduce the CAR gene. Although complex, this technique minimizes the risk of unrestrained genomic integration. The T-cells are then infused back into the patient.
The benefits of CAR-T-cell therapy include the MHC independent antigen recognition CAR-T cells provide. This lessens the impact of mechanisms used by tumor cells for immune escape such as the downregulation of MHC molecules. CAR-T-cells can also be used in patients regardless of their specific human leukocyte antigen (HLA) type. HLA protein markers are found on most cells in the body and allow the immune system to identify “self”. CAR-T-cells also allow for antigens other than proteins, like carbohydrates and lipids to be recognized.
Limitations to CAR-T-cell therapy include the identification of tumor specific target antigens. The binding of CARs is limited to molecules present on the surface of tumor cells. To select optimal antigens for targeting, the antigen needs to be selectively expressed on tumor cells at high levels but not be expressed on the surface of important normal tissue. Another limitation is the suboptimal performance of CAR-T cells in the treatment of solid tumors as it is difficult to find tumor specific antigens that are highly and uniformly expressed. Also, CAR-T-cells have the possibility of becoming dysfunctional due to a hostile tumor microenvironment characterized by oxidative stress, nutritional depletion, acidic pH or hypoxia. The presence of inhibitory soluble factors and cytokines and suppressive immune cells can contribute to T-cell inhibition.
Using CAR-T cells for the treatment of cancer can induce various side effects including high fever, low blood pressure, infection, low blood cell count or a weak immune system. A commonly observed toxicity includes cytokine release syndrome, which is characterized by high fever, hypotension, hypoxia, and multi-organ toxicity. CAR‐T cell related encephalopathy syndrome is another toxicity that results in a toxic encephalopathic state with symptoms of confusion, delirium, and occasionally seizures.
CAR-T cell therapy has had success in treating hematologic cancers like B‐cell malignancies, including acute and chronic B‐cell leukaemias, and B‐cell non‐Hodgkin lymphomas. Using anti-CD19 CAR-T-cells, there was a reported 90% complete remission rate in patients with B cell acute lymphoblastic leukemia. CD19 is antigen recognized as a target for immunotherapy in B cell malignancies because of its limited expression on mature B cells.
Immunotherapy remains a very active area of cancer research and scientists are constantly studying new ways to use immunotherapy in the treatment of cancer. PD-1/PD-L1 and CTLA-4 inhibitors along with CAR-T-cell based therapies are only the beginning of a new frontier of cancer treatment.