CAR-T CELL THERAPY (part3)
Control mechanisms
By adding a synthetic control mechanism to genetically engineered T cells, physicians can precisely control the persistence or activity of T cells in the patient's body to reduce toxic side effects. The main control techniques trigger T cell death or limit T cell activation and often regulate T cells via a separate drug that can be delivered or withheld as needed. [19]
Suicide genes: genetically engineered T cells are manipulated to contain one or more genes that can induce apoptosis when activated by an extracellular molecule. Herpes simplex virus thymidine kinase (HSV-TK) and inducible caspase 9 (iCasp9) are two types of suicide genes that have been integrated into CAR -T cells. In the iCasp9 system, the suicide gene complex consists of two elements: a mutant FK506-binding protein with high specificity for the small molecule rimiducid/AP1903 and a gene encoding a human caspase 9 with reduced pro-domain. Administration of rimiducid to the patient activates the suicide system, resulting in rapid apoptosis of the genetically modified T cells. Although both the HSV-TK and iCasp9 systems have shown remarkable function as safety switches in clinical trials, several shortcomings limit their application. HSV-TK is derived from viruses and may be immunogenic to humans. In addition, it is currently unclear whether suicide gene strategies act rapidly enough in all situations to stop dangerous off-tumor cytotoxicity. [20]
Dual antigen receptor: CAR -T cells are engineered to express two tumor-associated antigen receptors simultaneously, reducing the likelihood that T cells will target non-tumor cells. Reportedly, CAR -T cells with dual antigen receptor have less severe side effects. An in vivo study in mice shows that CAR -T cells with dual receptor effectively destroy prostate cancer and achieve complete long-term survival. [21]
ON switch: In this system, CAR -T cells can only function when both a tumor antigen and a benign exogenous molecule are present. To achieve this, the chimeric antigen receptor of the CAR -T cell is divided into two separate proteins that must come together to function. The first receptor protein usually contains the extracellular antigen-binding domain, while the second protein contains the downstream signaling elements and co-stimulatory molecules (such as CD3ζ and 4-1BB). In the presence of an exogenous molecule (e.g., a rapamycin analog), the binding and signaling proteins dimerize together, allowing the CAR -T cells to attack the tumor. [22]
Bispecific molecules as switches: bispecific molecules target both a tumor-associated antigen and the CD3 molecule on the surface of the T cells. This ensures that T cells can only be activated when they are in close proximity to a tumor cell. The anti-CD20/CD3 bispecific molecule shows high specificity for both malignant B cells and cancer cells in mice. [76] FITC is another bifunctional molecule used in this strategy. FITC can redirect and regulate the activity of FITC-specific CAR -T cells on tumor cells with folate receptors. [22]
2.6 CAR-T CELL THERAPY SIDE EFFECTS
2.6.1 Cytokıne release syndrome
Cytokine release syndrome reported in 54-91% of cases. After CAR -T-cell infusion proinflammatory cytokines. 1-2 after first infusion fever, tachycardia, hypotension, hypoxia that develops after hours, decrease in ejection fraction, renal failure, elevation of liver enzymes and coagulation disorders (prolongation of PT /APTT). In the treatment of standard supportive therapy and IL -6tocilizumab or corticosteroids with receptor antagonistsApplies. Corticosteroids, CAR -tocilizumab because it can suppress cellular activity, tocilizumab is the first immunosuppressant of choice. [12]
2.6.2 NEUROLOGICAL CHANGES
Cytokine release syndrome is reported in 54-91% of cases. After CAR -T-cell infusion proinflammatory cytokines. 1-2 after the first infusion fever, tachycardia, hypotension, hypoxia that develops after hours, decrease in ejection fraction, renal failure, elevation of liver enzymes and coagulation disorders (prolongation of PT /APTT). In the treatment of standard supportive therapy and IL -6tocilizumab or corticosteroids with receptor antagonistsApplies. Corticosteroids, CAR -tocilizumab because it can suppress cellular activity, tocilizumab is the first immunosuppressant of choice. [12]
2.6.3 B CELL APLASIA
An important side effect of anti-CD19/CD20 CAR-T cell therapy is B cell aplasia. [12]
2.7 CAR-T CELL THERAPY AND CLINICAL APPLICATIONS
2.7.1 ACUTE LYMPHOBLASTIC LEUKEMIA
Induction chemotherapy for acute lymphoblastic leukemia (ALL). Although the response rate is 80-90%, approximately 50% of patients develop relapse within 5 years. ALL In a phase II study of 61 pediatric patients with relapsed refractory leukemia ALL, the overall response rate was 20%, the median response time was 29 weeks, and the median survival time was only 13 weeks. The Phase I study II, which enrolled 70 pediatric patients with relapsed refractory ALL with the bispecific CD19-CD3 antibody blinatumomab, another treatment option, found a complete response of 39%, minimal residual disease negativity of 20% and median survival of only 7.5 weeks. ALL Most cancers, especially relapsed refractory B- ALL, are well suited for CAR -T cell therapy. CD19 is an ideal target for CAR -T therapy. However, CD20, immunoglobulin light chain, or CD22 are other potential targets listed at ALL because of the expected side effect of immunotherapy, "antigen escape." The National Cancer Institute (NCI), College of Pennsylvania (UPenn) and Memorial Sloan-Kettering Cancer Center (MSKCC) are conducting clinical trials using CART -19 second-generation cells at ALL. NCI and MSKCC used CD28 as the co-stimulatory domain, while UPenn preferred 4-1BB.19 75 children and young adults with CD19+ relapsed refractory B- ALL from a total of 25 centers in North America, Europe, Israel, and Australia. CAR T-cell therapy was administered in the multicenter Eliana trial, which enrolled patients aged 23 years. Patients included in this study had received a median of three prior treatment series, 61% had undergone allogeneic stem cell transplantation, and the median bone marrow blast rate was 74%. Patients were infused with a median of 3.1 x 106/kg CART -19 cells. Overall survival at 12 months was 76% and event-free survival was 50%. Grade 3-4 adverse events were observed in 73% of cases. Cytokine release syndrome was seen in 77% (tocilizumab was administered in 48% of patients) and neurologic events were seen in 40% of patients. With these study results, tisagenlecleucel became the first gene therapy product approved in the U.S. for the treatment of refractory or second or subsequent relapsed B-cell progenitors ALL under 25 years of age. [23]
2.7.2 LYMPHOMA
The prognosis of patients with lymphoma who do not respond to primary or salvage therapy or who relapse after autologous stem cell transplantation is extremely poor. In the recently published SCHOLAR-SCR, which enrolled 636 patients with refractory diffuse large B-cell lymphoma (DLBCL) (refractory to chemotherapy or relapse ≤ 12 months after autologous stem cell transplant)
In the first study, the overall response rate was 26%, the complete response rate was 7%, and the median survival was only 6.3 months. The first study showing the efficacy of anti-CD19 CAR -T cells in DLBCL was organized by the NCI. Eleven patients with different subtypes of B-cell NHL (DLBCL: 4, primary mediastinal B-cell lymphoma: 4, DLBCL transformed from CLL: 1, indolent NHL not elsewhere classified:1, splenic marginal zone lymphoma:1) CAR -T cells were infused once after the combination of fludarabine-cyclophosphamide chemotherapy. Response was achieved in 8 of 9 patients whose response could be assessed, and complete response was achieved in 5 of these patients. In 32 patients (aggressive B-cell lymphoma: 11, transformed follicular lymphoma: 11, follicular lymphoma: 4, mantle cell lymphoma: 4), the co-stimulatory domain was preferred, and overall response was 63% and complete response was 33%.23 In another study published by the NCI, 22 patients with different subtypes of B-cell NHL (DBCL: 13, transformed follicular lymphoma: 4, follicular lymphoma: 2, primary mediastinal B-cell lymphoma: 2, mantle cell lymphoma: 1) were administered 19- 28 CAR -T cells at a dose of 1, 2, or 6x106/kg after the combination of cyclophosphamide-fludarabine chemotherapy. While overall response was 73% and complete response was 55%, neurological adverse events occurred in 55% of patients. In this study, a low-dose cyclophosphamide-fludarabine combination was shown to be sufficient to decrease lymphocyte counts and increase serum cytokine levels. [23,24]
In the treatment of lymphoma, in addition to CD19, other antigens such as CD20, CD30 and the light kappa chain are also targeted. There are two important reasons for this. First, Hodgkin lymphomas, T-cell lymphomas, and even some B-cell NHLs may not express CD20. Second, CD19 may not be expressed in all cells prior to treatment or may be lost during treatment due to persistence or redevelopment of resistant clones. In this case, anti-CD19 CAR -T cell therapy may fail to treat CD19+ lymphomas. Therefore, researchers are developing CARs that target alternative lymphoma-associated antigens. [24]
2.7.3 CHRONIC LYMPHOCYTIC LEUKEMIA
The algorithms for CLL treatment have changed with the new agents that have been used recently. The B-cell receptor pathway inhibitors (BCRi) ibrutinib and idelalisib and the BCL2 antagonist (BCL2a) venetoclax are promising new agents for the treatment of CLL. Although high overall response rates are achieved in patients with relapsed/refractory CLL by using the new agents in monotherapy or combination therapy (with chemotherapy and/or monoclonal antibodies), drug resistance may develop, complete remission is rare with these agents, there is no curative potential, and their effect in Richter transformation is limited. In addition, TP53 abnormalities remain a poor prognostic factor in patients with relapsed/refractory CLL treated with BCRi. Long-term effects and side effects, the presence of indicators to predict efficacy, optimal treatment duration, and treatment management in patients who relapse during treatment with these agents are the unknown points of the new agents. Studies of the use of second- and third-generation CART -19 cells and CD28 or 4-1BB as co-stimulatory domains in CLL are ongoing. However, the early immunodeficiency caused by the pathogenesis of CLL negatively affects T-cell expansion and the in vivo proliferative response in these patients, limiting CAR -T-cell therapy. However, in a study by Fraietta et al, ibrutinib was shown to increase the expansion and antitumor effect of CART -19 cells. [25]
2.7.4 MULTIPLE MYELOM
Multiple myeloma (MM) remains an incurable disease associated with high mortality and morbidity. The successes achieved with CAR -T cells in leukemia and lymphoma have also spurred the development of CAR -T therapies for MM that require new treatment approaches. The most important factor in the success of CAR -T cell therapy is antigen selection. Several subclones have evolved in MM. They are responsible for the genetic and phenotypic heterogeneity of myeloma cells from the same patient, resulting in differences in cell surface antigen expression. While it is strongly and uniformly expressed on all malignant plasma cells, there is no known plasma cell antigen that is not expressed at all in normal cells. Antigens studied for anti-myeloma CAR -T cells: CD44 variant 6, CD70, CD56, CD38, CD138, CD19, immunoglobulin kappa light chain, SLAMF7 (signaling lymphocyte-activating molecule F7), and BCMA (Bcell maturation antigen). The first clinical trial (3 MM ) was conducted by the NCI and was focused on BCMA. [26]
In the study in which the patient participated, increasing doses of anti BCMA CAR -T cells with CD28 as a co-stimulator domain were administered after the cyclophosphamide-fludarabine chemotherapy combination (cyclophosphamide 300mg/m2, 3 doses; fludarabine 30mg/m2, 3 doses). Excellent complete response was achieved in 1 patient, very good partial response in 2 patients, partial response in 1 patient, and stable disease in 8 patients. While overall response was 73% and complete response was 55%, neurological adverse events occurred in 55% of patients. [26]
2.7.5 SOLID TUMORS
Targeting solid tumors with CAR -T cells is more difficult than targeting hematologic cancers. This is because solid tumors may have disruption of antigen expression or lack of antigen-presenting mechanisms due to the genetic instability of tumor cells. Other factors limiting the success of CAR -T-cell therapy in solid tumors include the histopathologic features of the tumor, inadequate delivery of CAR -T-cells to the tumor, a strong local immunosuppressive microenvironment, tumor heterogeneity, and lack of specific antigens.34 Although CAR T-cell treatment in solid tumors is promising, its results do not approach those of CD19 CAR T-cells. Targeted antigens include EGFR (epidermal growth factor), EGFRvIII, HER2 (human epidermal growth factor receptor), CEA (carcinoembryonic antigen), GD2 (disialoganglioside 2) for neuroblastoma; MSLN (mesothelin), PSMA (prostate specific membrane antigen), IL13Ra2 for glioblastoma; for squamous cell carcinoma of the head and neck, ErbB, VEGFR and FAP are included. [27]
2.8 CAR-T CELL THERAPY AND IMPROVEMENT STUDIES
Studies are ongoing on CAR designs developed to improve the current limitations of CAR-T cell therapy. By modifying the CAR design, a less immunogenic single-chain variable fragment (scFv) can be engineered to develop different co-stimulatory domains, optimize the spacer region of the receptor, and expand the CAR target antigen repertoire (e.g., CD20, CD22, CD30, and kappa light chain). targets. With advances in vector design, apoptosis genes will be incorporated to increase safety and genome editing technologies (CRISPR/Cas9) will be used to increase efficiency and reduce side effects. Production of cell products with a well-defined CD4+:CD8+ T cell ratio, production of less differentiated memory T cell subsets will be targeted. Efforts will be made to optimize CAR -T cell activity and reduce toxicity through changes in preparation regimens. Efforts will be made to increase CAR -T cell activity by adding pharmacological agents after cell infusion, e.g., infusion of immunostimulatory cytokines such as PD1/ PD -L1 inhibitors, ibrutinib, IL -15 {Tablo 2). [28]
