Recognizing and managing on toxicities in cancer immunotherapy

Abstract

Over the past 4 years, cancer immunotherapy has significantly prolonged survival time of patients with prostate cancer, melanoma, lung cancer, and liver cancer, but its side effects are also impressive. Different types of the immune therapeutic agents have different on-target or off-target toxicity due to high affinity or weak specificity, respectively. Treatment toxicity spectrums vary greatly even in patients with the same type of cancer. Common toxicities are fevers, chills, diarrhea colitis, maculopapular rash, hepatitis, and hormone gland disorder; therefore, routine monitoring of thyroid function, liver function, renal function, and complete blood count are absolutely necessary once treatment begins. Some side effects are reversible, and can be processed through the standard medicines. However, serious toxicities are lethal, which should be frequently followed-up, identified at an early stage and immediately symptomatic treated by high-dose immunosuppressors. In this case, thereafter, the same agent should not be challenged again.

Introduction

It has been more than 100 years since William Coley and his colleagues began cancer immunotherapy through intratumoral injections of live Streptococcus or inactivated Streptococcus pyogenes and Serratia marcescens, based on the observation that some patients with a bacterial infection had more extensive tumor regression compared with uninfected individuals. ¹ However, Provenge (sipuleucel-T), the first validation of modern immunotherapy vaccine, approved by the Food and Drug Administration (FDA) in April 2010 for advanced hormone-refractory prostate cancer, eliminated the disappointment of cancer immunotherapy. ² Recently, several emerging immunological approaches have significantly prolonged the survival time or improved quality of life for many cancer patients, such as renal cancer, melanoma, lung cancer, and liver cancer, and become one of the most potential treatments. ² Meanwhile, their toxic reactions that virtually affect every organ system should not fail to notice. Different types of cancer immune treatments are accompanied by special drug toxicities and variant side reaction spectrum, which are thought to be resulted from cross-interaction of multiple pathways. Off-target toxicities are generally unpredictable, as other unexpected targets are inhibited, which shared structures or epitopes with the expected targets. Common toxicities include increased capillary permeability, mild autoimmune response to normal tissues, and the autoinflammatory response which induced by cytokines, vaccine, and protein inhibitors, respectively. ³ Cytokines seem to likely produce a more diffuse non-specific T cell response; in contrast, checkpoint protein inhibitors, vaccines, and adoptive cellular transfer therapy seem to induce a more specific T cell activation. Activated T cell reacts directly with normal tissue cells, resulting in damage to specific organs. Severe toxicities to important organs need to be vigilant and identified at an early stage, and controlled by symptomatic treatment immediately. ⁴ Thereafter, the same agent should not be challenged again.

The cytotoxicities of cytokines

Interferon

In the 1980s, interferon-a (IFN-a) had been demonstrated to play a pivotal role in the course of antitumor activity in hairy cell leukemia, melanoma, renal cell carcinoma (RCC), and other solid tumors. Four years later, recombinant human IFN-a2 (Intron-A (Merck, Whitehouse Station, NJ); Roferon-A (Roche, Nutley, NJ)) was approved as a therapy for hairy cell leukemia, and in 1995, it became the first immunotherapy approved as the adjuvant treatment for the stage IIB/III of melanoma after treating with the FDA. ⁵

Contrary to initial expectations, IFN has not proven to be an innocuous therapy, despite it being a natural substance. The most common systemic toxicities of IFN therapy were fatigue and fever (ranged from 60% to 70%) followed by headache and myalgia, which were known as flu-like symptoms, in spite of affecting numerous organ systems. ⁶ Based on data from 143 patients treated with high-dose IFN-a in Eastern Cooperative Oncology Group (ECOG) Trial E1684, 66 patients experienced nausea and vomiting after IFN, 2/3 of the patients experienced early satiety, anorexia, and weight loss. These symptoms can be controlled by non-steroidal anti-inflammatory drugs, and severe fatigue often needs to decrease the dosage of IFN. Patients with fever >39°C more than 1 day should be assessed, because it may indicate systemic infection, especially who complained about cough, dyspnea, or respiratory symptoms should undergo radiologic evaluation. ⁷

Myelosuppression is the main adverse effect of dose-limited as high-dose interferon-a-2b inhibits the proliferation and activation of megakaryocytic progenitor cells in a non-lineage specific manner. Although neutropenia is reported in 65% patients (92/143) treated with high-dose IFN-a in E1684 trial, IFN does not result into neutropenic fever or sepsis and is rarely a cause for discontinuation of therapy. Thrombocytopenia and anemia were observed in about 10% of IFN therapy patients. Myelosuppression could be controlled by suspending or reducing the dose, but anyway, complete blood count should be monitored weekly during induction phase, monthly during the first 3 months of maintenance phase, and every 3 months thereafter. Thrombocytopenic purpura and progressive anemia are rare, but permanent withdrawal from IFN is required.^8,9

In a preliminary study of IFN adjuvant therapy for high-risk melanoma patients, hepatotoxicity resulting in two deaths was observed. ¹⁰ Liver function should be assessed at baseline, especially in the presence of hepatitis B or C, weekly during induction, monthly during the first 3 months of maintenance, and then every 3 months during the remainder therapy, although mild elevations in hepatic enzymes without clinical symptoms are common. It is widely considered that IFN should be suspended in patients with grade 3 liver toxicity, until the transaminase concentration returned to within 1.5 times of normal level, and if reused, IFN should be reduced by at least 30%.

Some side effects from IFN are completely independent of dose and duration, such as autoimmune diseases, a biomarker related to good prognosis. There is a prevalence of 10% to 15% of thyroid dysfunction (hyperthyroidism or hypothyroidism) in patients treated with IFN. ¹¹ After a long period of hyperthyroidism, hypothyroidism usually appears, thus T3 and T4 levels, and thyroid autoantibodies should be measured in all patients. The rare Sarcoidosis is most likely to have an autoimmune basis and presents skin lesions similar to subcutaneous metastases or mediastinal lymph nodes with Fluorine-18 deoxyglucose (FDG) high-uptake in positron emission tomography. ¹² Also, vitiligo, lupus, rheumatoid arthritis, polymyalgia rheumatica, and psoriasis are reported. In patients with pre-existing autoimmune disease, IFN should be used with caution, because the following IFN therapy generally induces more serious side effects, possibly involved in the memory function of immunity system. ¹³

The most common neuropsychiatric side effects are depression and irritability with a frequency of as many as 24% patients; however, an acute confusional state, anhedonia, fatigue, apathy, sleep disturbances, sexual dysfunction, impaired memory, cognitive dysfunction, and suicidal ideation induced by IFN-a are also reported. Neuropsychiatric symptoms generally are mild to moderate and resolve within 2–3 weeks of IFN discontinuation. ¹⁴ However, in some patients, neuropsychiatric toxicities can persist for months or even years. Antidepressants appear to be effective. For example, paroxetine is prophylactically administered to 20 patients with malignant melanoma, resulting in decreased symptoms of major depression with 11% of aroxetine-treated patients versus 45% of placebo patients. Patients with history of severe depression are prohibited from IFN, although prophylactic use of antidepressant drugs is able to reduce the risk of depression. Nevertheless it is also notable that the use of antidepressants may worsen depression symptom. ¹⁵

Interleukin-2

Interleukin-2 (IL-2), a glycoprotein first described in 1976 as a T-cell growth factor, plays a central role in immune regulation and T-cell proliferation. In 1998, high-dose cytokine of IL-2 was approved by the FDA based on its long-lasting antitumor responses in patients with advanced RCC or melanoma. High dose IL-2 should be administered under the supervision of the qualified physicians well versed in anticancer agents. ¹⁶ An intensive care facility and specialists skilled in cardiopulmonary monitoring or hemodynamic support are necessary to treat severe toxicities of IL-2 such as pulmonary edema and hypotension . The rate of drug-related deaths in metastatic RCC patients who received single-agent IL-2 was 4% (11/255) and 2% (6/270) in metastatic melanoma patients.

IL-2 common toxicities include chills, fever and fatigue, nausea, vomiting, diarrhea, hypotension, elevated transaminases, dyspnea, hyperbilirubinemia, and oliguria. ¹⁷ Capillary leak syndrome (CLS) begins immediately after treatment with IL-2 at the recommended dosages, which is characterized by a loss of vascular tone and extravasation of plasma proteins and fluid into the extravascular space. ¹⁸ CLS can lead to hypotension, and the reduction of organ perfusion might be too severe to result in death. CLS may be associated with cardiac arrhythmias (supraventricular and ventricular), angina, myocardial infarction, and pleural effusion, as well as occasional pulmonary edema, gastrointestinal bleeding or infarction, prerenal azotemia, and mental status changes. Hence, IL-2 should be restricted to patients who hold with normal cardiac and pulmonary functions. Extreme caution should be given to patients who were found to have a history of cardiac or pulmonary disease by means of normal thallium stress test and normal pulmonary function test. Hypotension generally is dose-related, and can be managed through vasopressors, even outside the intensive care unit. ¹⁹ With the possible development of extravascular fluid accumulation, ascites, pleural, or pericardial effusions, management of these events depends on a careful balance of fluid shifts so that neither the consequence of hypovolemia (e.g., impaired organ perfusion) nor the consequences of fluid accumulation (e.g., pulmonary edema) exceed the patient’s tolerance. Clinical experience has shown that early administration of dopamine (1–5 µg/kg/min) to patients manifesting CLS before the onset of hypotension can help to maintain organ perfusion especially to the kidney and thus preserve urine output. If organ perfusion and blood pressure are not sustained by dopamine therapy, clinical recommendations are to increase the dose of dopamine to 6–10 µg/kg/min or add phenylephrine hydrochloride (1–5 µg/kg/min) to low dose dopamine. ²⁰ Patients receiving vasopressors should be monitored by telemetric cardiac monitoring, as adrenergic agonists can induce atrial arrhythmia. In few patients, evidence of myocardial injury, including findings compatible with myocardial infarction or myocarditis, has been reported, which usually can be resolved within a few days, and without any sequelae. Therefore, patients should be surveilled through cardiac monitoring until cardiac enzymes are down to normal level, and the subsequent IL-2 therapy should be performed cautiously.

Thrombocytopenia, anemia, and abnormal coagulation function are observed in high-dose IL-2 treatment. Impaired neutrophil function (neutrophil chemotaxis) leads to an increased incidence of disseminated infection including sepsis, bacterial endocarditis, and catheter infection, ²⁰ which could be significantly reduced through prophylactic antibiotics with oxacillin, nafcillin, ciprofloxacin, or vancomycin.

Almost all adverse events (AEs) related to IL-2 might be controlled through suspension or cessation of administration. However, clinical trials have shown that autoimmunity, neurotoxicity, and myocarditis may worsen or continue for a period of time (mostly within 4 weeks) after IL-2 withdrawl.^16,21 The cause of long-term autoimmunity reactions is unknown. Autoimmune diseases, such as hypothyroidism, need over 6–10 months to recover; in contrast, vitiligo may deteriorate or even discontinue the therapy. The neurotoxicity of IL-2 is mild, including lethargy and irritability, or psychosis florid, which can be present as an episode of psychosis. Neurotoxicity reaches to the maximum within 24 h after the last administration, and should be vigilant and early recognized.

Glucocorticoids, rather than non-steroidal anti-inflammatories, should be avoided to concomitant administration with IL-2 due to reducing IL-2 potential benefit from antitumor effectiveness, although these agents have been shown to relieve IL-2-induced side effects including fever, hyperbilirubinemia, confusion, and dyspnea. Many agents have been developed to reduce the toxicities of IL-2 through blockade of its receptor; however, these antagonists were widely used in cancer patients so far because of their mild efficacy. ²²

Tumor necrosis factor

Since tumor necrosis factor (TNF) was found by Carswell in 1975, which acts as a pro-inflammatory and anti-inflammatory cytokine that was identified as a significant factor participating in the development of autoimmune disease and cancer, as well as the protection against infectious pathogens, ²³ numerous anti-TNF immunotherapy cases have been described. In fact, anti-TNF immunotherapy was synergic with apoptotic-cell-based therapy which is known as an efficient treatment with human cancer including colorectal cancer and breast cancer. ²⁴ Unfortunately, unexpected side effects have been described accompanying the anti-TNF therapy in clinical practice. The main toxicities of anti-TNF therapy include fever accompanied by hypotension and flu-like symptoms (chills, joints, and aches), as well as decrease of platelet and white blood cells but increase of transaminase.

Compelling evidence has reported that paradoxical adverse events (PAEs), including psoriasis, Crohn’s disease, and hidradenitis suppurativa, mainly emerge with anti-TNF-α agents. Other PAEs could be classified as borderline including uveitis, scleritis, sarcoidosis, and other granulomatous diseases (granuloma annulare, interstitial granulomatous dermatitis), vasculitis, vitiligo, and alopecia areata. Specially, it is found that the risk for paradoxical psoriasis is non-significant based on the data from registries. PAEs resulted by anti-TNF-α might be managed mainly depending on various aspects, such as the type and severity of the AEs, pre-existing treated conditions, and the possibility of alternative therapeutic options for the underlying disease. ²⁵

IL-12

IL-12 was previously called cytotoxic lymphocyte maturation factor (CLMF) or natural killer cell stimulatory factor (NKSF) composed of two subunits, which was identified as a powerful immunostimulatory cytokine with a strong antitumoural activity. It has been about 10 years since IL-12 was proved efficacious in numerous preclinical tumor immunotherapy studies. ²⁶ The study regarding comparisons of IL-12 and conventional chemotherapy in the treatment of lung cancer has revealed that IL-12 treatment suppressed lung tumor growth, resulting in the long-term survival of lung cancer–bearing mice. ²⁷ Furthermore, studies have indicated that IL-12 seems to be less toxic than other immunotherapeutics such as IFNs or IL-2. ²⁶

Despite encouraging immunostimulatory and antitumor effects of IL-12, IL-12-related toxicities should be taken into consideration. In serum liver function tests, the results suggested that common side effects mainly included fever, chills, fatigue, nausea or vomiting, and asymptomatic elevation, as well as transient neutropenia and thrombocytopenia. Fortunately, there was no patient who required platelet transfusion or had a neutropenic fever, indicating transient neutropenia and thrombocytopenia were insignificant. However, when three patients were treated with 250 ng/kg recombinant human IL-12, dose-limiting toxicities occurred in two of them, including diarrhea and asymptomatic elevation. ²⁸ Therefore, appropriate doses of IL-12 can be given safely to patients, especially to patients after autologous stem cell transplantation for high-risk hematological malignancies. Further examinations should be performed to assess the efficacy of IL-12 in the immunotherapy setting.

The toxicities of cancer vaccines

To date, although cancer vaccines have been available since 1970s, only one cancer therapy vaccine sipuleucel-T has been approved in view of increasing survival of patients with castration-resistant metastatic prostate cancer in two trials. Overall, cancer vaccines are well tolerated with predominant grade 1 toxicities including chills, fever, fatigue, back pain, nausea, erythema, and itching at the injection site, but over grade 3, AEs are also observed occasionally. ²⁹ Rahma et al. ³⁰ analyzed the toxicity profile of all 239 phase I, phase I/II, and pilot studies for therapeutic cancer vaccines conducted from 1990 to 2011, and found that the rate of grade 3/4 vaccine-related systemic toxicities was 1.25 AEs per 100 patients and 2 per 1,000 vaccines. Only two of the 127 dose-escalation trials reported vaccine-related dose-limiting toxicities, both of which used bacterial vector vaccines.

Cancer vaccine toxicities are associated with types of vaccines, rather than doses of vaccines. Data had shown that autologous vaccine trials had a toxicity rate of 1.36 events per 100 patients. Five toxicities are reported in the allogeneic and 35 in the synthetic vaccine trials, giving a vaccine-related toxicity rate of 1.23 events per 100 patients in each group. The highest vaccine-related toxicity rate among the synthetic vaccines is reported in the bacterial vector trials (3.97 events per 100 patients). In the 15 trials analyzing dose-related toxicity vaccine-related toxicities, only six reported the toxicities at the highest dose level. Two of these six trials that reported toxicities at the highest dose level used bacterial vaccines, and the rest used autologous, DNA, viral, or liposomal vaccines. Two toxicities occurred in the middle dose level in one trial that used a bacterial vaccine, with no further toxicity occurring when the dose was escalated. Four toxicities occurred at the lowest dose level in three trials, two with peptide vaccines and one with a liposomal vaccine. Interestingly, none of these trials reported further toxicities when the dose was increased.

Almost all of the cancer vaccines usually produce only low-level toxicities. The reason maybe is that many tumor-associated antigens are significantly overexpressed in cancer cells, but rarely expressed or undetectable in normal cells. ³¹ For example, most melanoma vaccines are directly against the differentiation antigen of melanoma cells, thus occasionally mild immune toxicities occur, such as vitiligo, which are associated with a good therapy response. Of course, the vast majority of cancer vaccines are tested in preclinical or phase I–II clinical trials with a small-size sample. ³² Most importantly, many cancer vaccine trials use the “3 + 3 design,” which may not be the most suitable approach to gathering dose-toxicity information from early phase trials, so that AEs are not able to be comprehensively and accurately assessed. ³³

Currently approved sipuleucel-T, a vaccine prepared by a 250-mL suspension containing a minimum of 50 million autologous CD54+ cells activated with PAP-GM-CSF (prostate acid phosphatase–granulocyte-macrophage colony-stimulating factor), has mild toxicity, such as transient chills, fatigue, and fever within the first 24 h after the injection. The most common AEs, reported in patients treated by sipuleucel-T at a rate ≥15%, are chills, fatigue, fever, back pain, nausea, joint ache, and headache. Grade 4 AEs (life-threatening) have an incidence rate of less than 4%.^34,35 In 95.1% of patients reporting acute infusion reactions, the events were mild or moderate, based on the analysis on 601 prostate cancer patients. Fevers and chills generally are resolved within 2 days (71.9% and 89.0%, respectively). Sipuleucel-T is discontinued in 1.5% of patients due to AEs. Some patients who required central venous catheters for treatment with sipuleucel-T developed infections, including sepsis. Monitoring for infectious sequelae in patients with central venous catheters is recommended. The mechanism of AEs may be that specific antigen encoded by prostate acid phosphatase fusion protein, selectively induced to specific immune response to cancer cells, has expression in the normal cells but peptide epitope is not high enough, resulting in weak immune response. ³⁶ The combination of antigen specific cancer vaccine and checkpoint inhibitors do not appear to lead to increased toxicity.

Although the low toxicity of cancer vaccines may reflect their corresponding with moderate immune activities, there are more than 400 open studies of cancer vaccines based on the antigens selection of peptide or protein, dendritic cells (DCs), or DNA recorded in ClinicalTrials.gov so far. ³⁷ These trials are testing vaccines across a wide range of cancers, including non-small cell lung cancer (NSCLC), melanoma, glioblastoma, triple negative breast cancer pancreatic adenocarcinoma, and prostate cancer. In addition, there are a range of strategies of vaccines combined with chemotherapy, radiotherapy, targeted therapy, and immune suppression antibodies, which may cause higher toxicities.

Toxicities caused by adoptive cell therapy

Adoptive cell infusion therapy (ACT) using naturally occurring tumor-reactive lymphocytes has been applied in patients with melanoma, liver cancer, pancreatic cancer, and renal cancer worldwide, especially in Europe and East Asia, although there were a series of contradictory opinions on its extensive clinic practices. ACT was performed through a multi-step procedure: first, the T cells were isolated from peripheral blood monocytes, tumor infiltration lymphocytes (TIL), or chimeric antigen receptors (CARs) of T cells that were genetically engineered by monoclonal antibody variable region; second, T cells were activated and amplified by immunostimulatory molecules in vitro; finally, sufficient T cells that possess not only the necessary functions to mediate cancer regression but also high tumor specificity were re-infused into patients. ³⁸ Some widely spread cancers can be effectively treated with ACT. Furthermore, the success rate of T-cell transformation can be improved by the combination chemotherapy of lymphodepleting preparative regimen before the cell infusion.

In general, ACT has not marked side reactions, because of the usage of autologous cells for infusion, such as cytokine-induced killer (CIK) cells, activated natural killer (NK) cells, or TIL cells. ³⁹ Although clinical practices are conducted under greatly various conditions, the AEs are uncommon chills, fever, and fatigue, all of which are no more than 4% and grade 2, at least in our cancer center. However, the fatigue, we thought, is partly associated with diphenhydramine which was used in pretreatment to prevent acute infusion reactions. Serious fever (temperature: >39°C) should be immediately managed by non-steroidal anti-inflammatory drugs like corticosteroids. Patients aged less than 50 years have a higher risk of chills followed by fever during administration of gamma delta T (γδ T) cells or NK cells, if not be premeditated orally with acetaminophen and an antihistamine. The underlying mechanism of this response is still unknown, but there is no doubt that younger patients have a stronger immunity response. Using ACT as the chemotherapy regimens prior to human cancer immunotherapy can lead to not only the loss of lymphocytes but also a decrease of both neutrophils and platelets for 7–10 days.^40,41 Although severe events are rare and can be controlled, and bone marrow function can soon recover from injury, the patients are at a risk of sepsis and bleeding. In the treatment of patients with lymphodepleting preparative regimens, sepsis is the leading cause of death, accounting for 1%–2%. ⁴²

However, adoptive T cells that are genetically engineered to express conventional T cell receptors (TCRs) or CARs are able to produce on-target and off-target toxicity triggered by anomalous immune activation. These AEs in some cases are fatal, which not only require high levels of professional personnel to deal with but also are great challenges faced by investigators.

Toxicities related to high affinity of TCR

Toxicities induced by antigens which are derived from tumor-restricted expression proteins are related to high affinity, rather specificity of TCRs, for which they are named as on-target toxicities. The marked potency of T cells through encoding higher affinity CARs enables the recognition of minute levels of antigen expressed on normal cells. Thus, bounding normal, non-mutated antigenic targets that are expressed on normal tissues but overexpressed on tumor cells have led to severe on-target, off-tumor toxicities. It is well known that potential cancer rejection antigens, such as MART-1 melanoma-melanocyte differentiation antigen or gp100, are formed by non-mutated protein highly expressed in malignant melanoma cells but rarely expressed in normal melanocytes. To identify these antigens, TCRs were engineered into retroviral vectors and used to transduce autologous peripheral lymphocytes administered to 36 patients with metastatic melanoma in a pilot study. ⁴³ As a non-surprising result, patients exhibited destruction of normal melanocytes in the skin, eye, and ear, due to fully activated T cells, with a strong ability to recognize cognate antigens of MART-1 and gp100 expressed in their melanocytes. And sometimes in patients with severe on-target and off-tumor toxicities, steroid administration was required to treat uveitis and hearing loss, ⁴⁴ fortunately which can be controlled by application of agents. Some factors, such as the expression level and the distribution of the attacked protein in normal tissues, and whether the affected cells immediately endanger the survival of the patient, determined the severity of damage from immunotherapy.

It is not beyond our expectation that higher affinity CARs also led to obvious AEs. Since effective clinical treatment with anti-CD19 CAR T cells was reported for the first time in 2010 after a patient with advanced-stage lymphoma treated at the National Cancer Institute (NCI) experienced a partial remission of lymphoma and long-term eradication of normal B cells, anti-CAR CD19 therapy for refractory lymphoma or leukemia has become acceptable. ⁴⁵ The CD19 antigen molecule expressed on more than 90% of B-cell malignancies at all stages of B cells differentiation but not on other normal cells. Data from an extended sample trial that enrolled 15 patients with diffuse large B-cell lymphoma, indolent lymphomas, and chronic lymphocytic leukemia showed that grade 3 and 4 toxicities experienced by patients are fever, hypotension, and infection, as well as neurologic toxicities, mostly occurred during the first 2 weeks after infusion. One patient who had chemotherapy-refractory primary mediastinal B-cell lymphoma (PMBL) with extensive fibrotic mediastinal lymphoma involvement died suddenly, without experiencing signs of cytokine-release toxicities such as fever. A likely cause of death is cardiac arrhythmia. A risk of infection that severe on-target toxicity B cells lack caused by T cells attack can be eliminated through intravenously injected immunoglobulin G or the IL-6 receptor-blocking antibody tocilizumab. ⁴⁶

When the carcinoembryonic antigen (CEA) was targeted by T cells in colorectal cancer, all three patients had a serious risk of life-threatening colitis; similarly, a low expression of CEA existed in normal colon tissue. ⁴⁷ The use of T cells with CAR specific to HER-2 may cause lethal lung injury, being manifested as acute pulmonary edema and hypoxia. The observation of death caused by acute lung toxicity indicated that it is probably because of the low-level expression of HER-2 antigen in mistakenly attacked lung epithelial cells. ⁴⁸ Some research groups have experimented to change amino acid sites by site-specific mutagenesis in TCR to enhance the affinity of TCR antigen binding. Carbon anhydride 9 is overexpressed in renal cell carcinoma patients, which is also expressed in bile duct epithelium, ⁴⁹ and it is not surprising that CAR T-cell therapy encoding anti-carbon anhydride 9 would lead to severe liver toxicity in clinical trials. When T-cell therapy leads to life-threatening toxicity (either cytokine release or autoimmunity), standard interventions are necessary, including the use of high doses of corticosteroids and alemtuzumab (an anti-CD52 antagonist), to inhibit or eliminate lymphocytes (which may lose all of the antitumor effects). A potential method is ongoing to look for TCRs or CARs against specific surface antigens expressed on tissues that are not essential for survival.

Toxicities related to weak specificity of TCR

On theoretical grounds, cancer rejection epitopes may be derived from two classes of antigens. The first class of cancer antigens is formed by non-mutated proteins with incomplete T-cell tolerance caused by their restricted tissue expression pattern. The second class of cancer antigens is formed by peptides that are derived from epigenetic mutated proteins in tumor self and thus entirely absent from the normal human genome, so-called neoantigens. ⁵⁰ Therefore, theoretically, toxicity induced by so-called neoantigens should not occur in normal tissues; unfortunately, diverse grades or types of AEs still were frequently reported. These unpredictable toxicity, related to weak specificity but high affinity of TCR or CAR, were primarily induced by a mechanism that the receptors on T cells identify cross antigen or epitope (or cross reactive) and then generate off-target and off-tumor toxicities associated with antigen independent activation. Almost certainly, TCR- or CAR-targeted non-mutated proteins also mismatched with non-aimed epitopes in normal tissues.

Melanoma-associated antigen A (MAGE-A) is a multigene family consisting of 12 homologous genes MAGE-A1 to MAGE-A12 located at chromosome Xq28 (11). MAGE-A3 is a cancer testis antigen not expressed in any of the normal tissues. Nine cancer patients were treated with adoptive cell therapy using autologous anti-MAGE-A3 TCR-engineered T cells. Three patients experienced mental status changes, and two patients lapsed into comas and subsequently died because of severe brain damage in white matter. The neurological toxicity observed in three cases was known to be that, T cells modified with an HLA-A*0201-restricted MAGE-A3-specific TCR recognize the different but similar epitopes of the MAGE-A12, which is expressed in human brain (and possibly MAGE-A1, MAGE-A8, and MAGE-A9) at quiet low level in the brain. ⁵¹ There are limited data that suggested toxicity was associated with T-cell dose, but it is likely to be variable in different individuals. Two patients (one multiple myeloma and another melanoma, respectively) treated with high-affinity TCR, which is specific to HLA-A1-limited MAGE-A3^a3a, experienced with the fatal cardiogenic shock, since the epitope derived from cardiac titin, ⁵² not related to MAGE-A3^a3a, is cross-recognized by the HLA-A1-restricted TCR. Therefore, strategies in order to enhance TCR affinity to neoantigens expressed in contracting normal key tissue and not be identified by typical preclinical screening analysis may lead to unanticipated toxicities. As these toxicities were difficult to relieve as soon as any essential normal organs represent potential targets, improved preclinical testing methods to better enable prediction of TCRs specificity have to be established in experimental models.

Cytokines release syndrome

One of the most common toxicity associated with the novel immunotherapies of CAR T and checkpoint inhibitors is cytokines release syndrome (CRS), a constellation of inflammatory symptoms resulting from cytokine elevations associated with T-cell engagement and proliferation. The patients experienced with CRS associated with T-cell therapies may represent a series of symptoms, including sepsis, fever, tachycardia, blood vessel leakage, less urine, and hypotension. ⁵³ Recently, multiple organ failures have emerged in severe cases and a CRS-related death after blinatumomab treatment has been reported. When treated with systemic IL-2 therapy, these symptoms appear more quickly; therefore, it suggests toxicities induced by cytokines releasing are directly mediated by IL-2. Without the use of IL-2, the onset of these symptoms may be delayed (T-cell infusion after 5–7 days). Even severe renal failure, coma, and respiratory failure can be completely reversed by supporting treatment. In a recent study of CAR CD19 in the treatment of leukemia, all patients had a certain degree of CRS, 27% of severe CRS need to maintain blood pressure treatment to compromise side effects of the cell therapy. ⁴⁶ IL-6 is an inflammatory cytokine involved in a large number of processes within the immune system, such as neutrophil trafficking, acute phase response, angiogenesis, B-cell differentiation, and autoantibody production. In a group of patients with B-cell malignancies treated by CAR modification of T cells (CD19 targeted), researchers found that IL-6 is a regulator of hemodynamic toxicity, and tocilizumab, a IL-6 receptor-blocking antibody, had obvious advantages to control early symptoms of CRS patients. ⁵⁴ Common treatment usually included intravenous fluid infusion, non-steroidal anti-inflammatory drugs, vasopressor drugs (if needed), and even cortisol hormones, and strong monitoring needs to be warranted.

Overall, compared with other types of immunotherapies, adoptive cell therapy is a more complex approach to cancer patients. Therefore, new technologies in improving the specificity of T cells should be further developed to do no or less damage to the deadly tissue in order to ensure the safety of ACT treatment. Second, lymphatic depletion dose of pretreatment should be carefully selected. Finally, further studies are needed to determine optimal toxicity surveillance and management, when serious AEs appear.

Some biomarkers to predict patients at high-risk of developing CRS related to immunotherapy, such as PRF1, MUNC13-4, STXBP2, and STX11, still necessitate new studies to establish. ⁵⁵ It has been advocated to integrate the suicide gene into the transplanted T cells, and this gene is enabled to be activated immediately once grade 3 or 4 AEs happened, ⁵⁶ but it is not clear whether it can be started in time to prevent permanent damage.

Toxicities in checkpoint protein inhibitors

Since 2011, FDA has approved three checkpoint protein inhibitory antibodies, including ipilimumab blocking cytotoxic T lymphocyte antigen-4 (CTLA-4) and pembrolizumab and nivolumab blocking programmed cell death protein-1 (PD-1), which have been used in the treatment of melanoma or NSCLC, and a variety of other forms of cancer. There are many agents tested now in clinical trials, such as tremelimumab (CTLA-4), MEDI4736 (programmed cell death ligand-1, PDL-1), and atezolizumab (PDL-1) developed in phase III. ⁵⁷ However, largely different to traditional cytotoxic agents with a narrow toxic spectrum and similar to targeted therapies, the toxicities (immune-related AEs (irAEs)) from each checkpoint protein inhibitor almost influence all the organs or tissues, thus which should not be overlooked, particularly in setting of combination therapy by concurrent or sequential methods. The most common clinically relevant toxicities included rashes, which may rarely progress to life-threatening toxic epidermal necrolysis, and colitis, characterized by a mild to moderate, but occasionally by severe and persistent diarrhea. ⁵⁸ The others with decreased incidence rates were hypophysitis, hepatitis, pancreatitis, iridocyclitis, lymphadenopathy, neuropathies, and nephritis. Therefore, at the beginning of treatment within the first 6 months, routinely monitoring of liver, renal, and lung functions, complete blood count is absolutely necessary per 6–12 weeks. If fatigue and non-specific symptoms are observed, hormone gland functions (especially the thyroid, pituitary, and adrenal glands) should be checked.^59,60 It is a noteworthy inflammation of the brain in patients with signs and symptoms of encephalitis including headache, confusion, and seizures also needed to be monitored. The serious toxicity should be considered for intravenous corticosteroids, and the frequency of inspection and follow-up should be increased, although some side effects were reversible and able to be dealt with in the clinic.

Organ-special toxicities of checkpoint protein inhibitors

One of the most common irAEs in patients who accepted checkpoint protein inhibitors treatment was the liver toxicity with elevations in serum levels of hepatic enzymes. Of 298 patients treated with ipilimumab, 39 cases had grade 3–4 toxicities. There was an increased incidence of liver test abnormalities in the melanoma group treated by nivolumab, with increases in aspartate aminotransferase (AST) (28%), alkaline phosphatase (22%), alanine aminotransferase (ALT) (16%), and total bilirubin (9%). ⁶¹ Grade 3–4 liver function abnormalities may require prolonged treatments, and it was appropriate to stop checkpoint protein antibodies and use high doses of cortisol, although no prospective trials have been conducted to specifically test whether one management strategy is superior. Standard precautions were that inhibitors have to be permanently discontinued for severe (grade 3) or life-threatening (grade 4) immune-mediated hepatitis. As among four patients for whom nivolumab in combination with ipilimumab was restarted, three had recurrence or worsening of hepatitis and one improved on corticosteroids.

In patients treated with ipilimumab, grade 3–4 colitis incidence rate is of 6%–14%, but which in great majority of PD-1/PDL-1 antibodies is less than 1%. However, diarrhea or colitis occurred in 21% (57/268) of melanoma and in 17% (50/287) of NSCLC receiving nivolumab. Colitis usually affects the distal colon and rectum, manifested as erythema, edema, erosion, and bleeding. In severe cases of patients who are hospitalized, microscopic examination revealed infiltration of neutrophils, and crypt damage or crypt microabscess. ⁶² Colitis usually occurs within 4–6 weeks in patients treated with PD-1/PDL-1 blockers, and in severe cases, the risk of colon perforation and obstruction (may also include the small intestine) may exist. ⁶³ The recovery period mostly was within 6 weeks, but extended in selected patients. Hospitalization should be considered for intravenous high-dose corticosteroids if symptoms were severe or refractory to oral corticosteroids therapy. Infliximab should be used unless symptoms could be cured or not recurrent after 3-day high dose glucocorticoid treatment. ⁶⁴ If intestinal perforation was suspected or diagnosed, administration of steroid hormones and surgery intervention should be considered. Infliximab is contraindicated to intestinal perforation and sepsis. There is no evidence that preventive oral steroid budesonide can prevent diarrhea occurrence. Physicians should always pay attention to the result of stool culture in order to exclude the infection due to deterioration of colitis.

Another common toxicity that occurred in ipilimumab therapy was skin side effect, and in 40% of patients, 2% are grade 3 or grade 4 toxicity. ⁶⁵ Common skin toxicities related to nivolumab also had a high prevalence ranged from 28% to 36%, although very few cases had three or four toxicities. These predominant dermatologic toxicities included maculopapular, erythematous rash and pruritus, but rare vitiligo and hair depigmentation were also reported, which were considered a positive prognostic factor in patients with melanoma. ⁶⁶ Mild skin toxicity might be relieved through using emollients, steroid cream (1% hydrocortisone), and antihistamines. Immune treatment needs to be terminated for grade 3 or 4 toxicity presented as Stevens Johnson syndrome and toxic epidermal necrolysis, which also should be evaluated by the dermatologists and treated with systemic high-dose corticosteroids (1–2 mg/kg/d prednisolone or corresponding drugs) followed by corticosteroid taper.

Almost no significant pneumonia was found in anti-CTLA-4 antibodies compared to anti-PDL-1 and PD-1 agents, suggesting that the spectrum of irAEs varies among different blocking antibodies. Grade 2–3 pneumonia resulted by nivolumab in NSCLC patients is 7%, but only 2.4% by pembrolizumab. Ipilimumab rarely causes pneumonia with symptoms with around 1% incidence rate. However, PD-1 antibody to treat patients may produce immune-mediated pneumonitis or interstitial lung disease identified as diffuse lymphocytic infiltration, and shortness of breath, sputum increasing, fever, chest pain, hemoptysis. The most frequent serious adverse reactions reported in at least 2% of patients receiving nivolumab were pulmonary embolism, dyspnea, pleural effusion, and respiratory failure. ⁶⁷ Seven deaths were due to pneumonia, pulmonary embolism, and limbic encephalitis. High dose glucocorticoid can result into remission of symptoms in most patients, but the recovery process might be prolonged.

The peripheral motor neuropathy and immune-mediated encephalitis are rarely found in patients treated with checkpoint protein inhibitors. Only <1% of patients were found to suffer from encephalitis in 8490 patients who received the treatments with nivolumab or combined with ipilimumab. ⁶⁸ However, the application of nivolumab is prohibited for patients diagnosed with moderate to severe neurologic symptoms or signs. ⁶⁹ Besides, the evaluation of treatment with nivolumab might include brain magnetic resonance imaging (MRI), lumbar puncture, and consultation with a neurologist, which is performed without the influences of infectious or other causes of moderate to severe neurologic deterioration. After the exclusion of other etiologies, corticosteroids are managed at a dose of 1–2 mg/kg/day prednisone equivalents for patients with immune-mediated encephalitis, followed by corticosteroid taper. Above all in the patients with immune-mediated encephalitis, the checkpoint protein inhibitors were permanently prohibited.

Severe infusion reactions have been reported in <1% of patients in clinical trials of checkpoint inhibitors as a single agent. ⁷⁰ It was recommended to discontinue treatment for patients with severe or life-threatening infusion reactions and interrupt or slow the rate of infusion for those with mild or moderate infusion reactions.

If ipilimumab trial is performed, patients with autoimmune disease or a history of viral hepatitis should be ruled out, but the latest data suggest that ipilimumab can be safely used in these patients. ⁷¹ Therefore, for the patients who recently suffered or are suffering from autoimmune diseases and have any types of inflammatory bowel disease, treatment should be carefully carried out.

Pharmacokinetic relationships of toxicities in the checkpoint protein inhibitors

The on-target or off-target toxicity caused by ipilimumab blocking CTLA-4 is related to does and efficacy. In case of administering dosage of ipilimumab from 3 mg/kg to 10 mg/kg, the rate of severe AEs associated with grade 3–4 increased from 5% to 18%, while the rate of serious AEs in 0.3 mg/kg was 0%. ⁷² However, 595 patients received 2507 doses of ipilimumab infused at either 3 mg/kg (n = 457) or 10 mg/kg (n = 138) over 90 min, and the 10-mg/kg group had no statistically significant higher incidence of infusion-related reactions than the 3 mg/kg group (4.3%vs 2.2%, P = 0.22). Similarly, out of 120 patients treated prospectively with ipilimumab 3 mg/kg infused over 30 min, seven patients (5.8%) had infusion-related reactions (p = 0.06, compared with 90-min infusions). ⁷³

For the anti-PD-1 antibody pembrolizumab, compared to 2 mg/kg (FDA-approved dose), patients who managed with 10 mg/kg every 3 weeks have much higher grade 3–4 AE. Recently, in a phase II clinical study for pembrolizumab, three or four drug-related AEs occurred in 12% patients, 5% of patients had severe AEs, and 3% of patients had to discontinue treatment because of AEs, although no drug-related deaths were reported. ⁷⁴ The most common AEs in all the grades included fatigue, pruritus, and rash. Anti-PD-1 antibody pembrolizumab and nivolumab seemed to be very similar in AE spectrum.

In contrast, the PD-1 antagonist nivolumab toxicity was similar in the different dose ranges of 0.3 mg/kg–10 mg/kg. In a randomized phase II trial, ⁷⁵ a total of 168 patients with metastatic renal cell carcinoma were randomly assigned to the different dosages of nivolumab treatment. The incidence of treatment-related AEs of any grade was similar across dose arms: 75%, 67%, and 78% in the 0.3-, 2-, and 10-mg/kg groups, respectively. The most common treatment-related AE is fatigue (24%, 22%, and 35%, respectively). In another study of 281 patients with melanoma, renal cell carcinoma, and lung cancer, patients were treated with doses of 0.3 mg/kg–10 mg/kg nivolumab, and 5% of patients experienced grade 3–4 irAEs. In a phase III trial ⁷⁶ of patients with untreated stage IV melanoma dosed by 3 mg/kg (FDA-approved dosage) nivolumab treatment, 11.7% have prevalence of grade 3–4 drug-related AEs, 6.8% suspend treatment as AEs, and 1% reported both impaired liver function and diarrhea.

Evidence has suggested that different types of cancer patients had different toxicities in PD-1 inhibitors trials. Immune-mediated hepatitis occurred in 1.1% (3/268) of melanoma patients receiving nivolumab. The time to onset of hepatitis in three patients was 97, 113, and 86 days after initiation of nivolumab. In NSCLC trial enrolled with 292 patients, however, only 0.3% patient developed immune-mediated hepatitis after 7.8 months of nivolumab exposure, resulting in permanent discontinuation. ⁶⁸ All patients received high-dose corticosteroids (at least 40 mg prednisone equivalents). Liver function tests improved to grade 1 within 4–15 days of initiation of corticosteroids. Among the patients with Hodgkin’s lymphoma who received nivolumab, 23 (22%) had grade 3 toxicity, including the most common skin rash, thrombocytopenia, fatigue, and fever, and two had transfusion reaction event for which monitoring was required. ⁷⁷ The prevalence of pneumonia caused by PD-1/PD-L1 antibody in melanoma was similar to that in NSCLC (2.9–3.4% and 3.4–3.5%, respectively), suggesting that the pre-existing lung disease may not contribute to the formation of injury. ⁷⁸ There was no evidence that toxicity profiles were associated with anti-nivolumab antibody development.

Although anti-CTLA-4 and anti-PD-1/PD-L1 antibodies have not been directly compared, immune-related effects associated with anti-CTLA-4 drugs were more common and of higher grade. ⁷⁹ Thus, patients experienced grade 3/4 adverse effects induced by anti-PD-1/PDL-1 antibodies are approximately 10%, which is lower than 20%–50% by anti-CTLA-4 antibodies. Compared to ipilimumab, however, anti-PD-1/PDL-1 therapy toxicity may need to receive long-term treatment for resolution of symptoms, and long-term monitoring. The irAE frequencies of PDL-1 antibody were lower than that of PD-1 inhibitors. In a study of a total of 207 patients who received PDL-1 antibody, the overall grade 3–4 toxicities incidence rate is 9%. Retrospective data indicated that corticosteroid treatment was not associated with impaired progression-free survival or overall survival. ⁸⁰

Toxicities in the checkpoint protein inhibitors-based combined therapy

Several studies have demonstrated that the irAE spectrum and various frequencies depended on the combination patterns of different agents. Generally, toxicities in combined therapy were more severe than those in agents alone. For instance, infusion reactions occurred in 1% (3/287) of patients receiving nivolumab and in 3% (3/94) of patients receiving nivolumab with ipilimumab. ⁸¹ In combination with nivolumab and ipilimumab, the irAEs of liver or small intestine and colon were common toxicities; moreover, the incidence of toxic reactions of grade 3 or 4 was up to 8%–10%, although the response rate was also improved to 43% and 53% cases had longer response duration. In a double-blind study involving 142 patients with metastatic melanoma who had not previously received treatment, drug-related AEs of grade 3 or 4 were reported more frequently in the combination group than in the ipilimumab monotherapy group (54% vs 24%) The most common grade 3 or 4 AEs associated with the combination therapy were colitis (17%), diarrhea (11%), and an elevated ALT level (11%). When ipilimumab combined with bevacizumab, the observed AEs were generally higher than expected with ipilimumab alone, but remained manageable. Importantly, there did not appear to be an increased incidence of dermatologic or gastrointestinal side effects such as colitis which are most concerning for ipilimumab. ⁸² In a randomized clinical trial with CTLA-4 blockade by ipilimumab and granulocyte-macrophage colony-stimulating factor (GM-CSF) combination, interestingly, grade 3–5 AEs occurred in 44.9% (95% CI; 35.8%, 54.4%) of GM-CSF plus ipilimumab and 58.3% (95% CI; 49.0%, 67.2%) of ipilimumab alone (two-sided p = 0.04). ⁸³

As with any combination therapy, the combination of checkpoint blockade immunotherapy and radiation contributes to the incidence of high-grade complications. For example, the immune-related side-effects of the specific site of irradiation include increased pneumonitis with lung irradiation, increased colitis with bowel irradiation and increased hypophysitis with central nervous system (CNS). ⁸⁴ Despite these side effects in patients, evidences demonstrated that the combination of radiation and checkpoint blockade immunotherapy can be safe for certain disease sites. In pelvis, as an example, the notable colitis was not found under the treatment in the combination of ipilimumab and radiation. Furthermore, favorable tolerance was obtained when ipilimumab in combination of stereotactic radiosurgery was applied for brain metastases. However, these results may be confused due to the limited samples. Therefore, to further evaluate the risks and toxicities caused by the combination of checkpoint blockade immunotherapy and radiation, it is urgent to carry out the additional phase I and II trials by incorporating safe endpoints in the definitive or palliative setting. ⁸⁵

However, it has to be emphasized that the grade 3/4 AE rate greatly varies from 0% to 66% (the median 21%) in a systematic review for 50 clinical trials with a total of 5071 patients receiving immune checkpoint inhibitors. ⁸⁶ Furthermore, around one fifth of the articles suggested that toxicities do not persist or are readily manageable. However, this claim is without presenting any data or evidence to support. Such trials that included incomplete data collection may significantly lead to a skewed representation of the tolerability of a novel immune checkpoint inhibitor.

Conclusions

Current immune agents exhibit a broad spectrum of toxicities such as vascular, dermatologic, endocrine, coagulation, immunologic, ocular, and pulmonary toxicities, instead of myelosuppression. And some immune-mediated adverse reactions may sustain for several months even after infusion is discontinued, which is obviously different to chemotherapy. The key to the successful prevention and management of immune therapy was early diagnosis, high vigilance, and good communication between physician and patients, or their caregivers, as well as rapid and active concurrent application of corticosteroids and other immunosuppressive agents. A better understanding of the mechanisms of action of these drugs and their interaction with the immune system was the foundation, based on that the new treatment strategies to improve A better understanding of the action mechanisms of these drugs and their interaction with the immune system provides the basis for the new treatment strategies for ACT improvement, which are involved in the identification and development of more specific antitumor T cells with the properties optimal for tumor regression. ⁸⁷

To date, there are no convinced biomarkers to predict immunotherapy toxicity across multiple tumor types, although it seems that patients with PD-L1 expression and also mismatch repair-proficient tumors have higher response rates to PD-1/PD-L1 blockade. It is thus hoped that advanced computing technologies and bioinformatics-based models can be innovated to predict adverse effects and identify and understand new targets as well as drug interactions, which are aiming to efficiently decrease rare or unexpected treatment-related adverse effects in the near future. ⁸⁸

Although cancer care represents a small fraction of overall health care costs, its contribution to health care cost escalation is increasing faster than those of most other areas. It is greatly considered that the introduction of new and costly drugs is an inescapable culprit. Thus, the term “financial toxicity” advanced by the American Society of Clinical Oncology (ASCO) has entered the vernacular as a means of describing the financial distress that now often accompanies cancer treatment. ⁸⁹ Financial toxicity, like organ toxicity, can also reduce quality of life and impede delivery of high-quality care, and is an important parameter of cancer therapy value to stratify the magnitude of clinical benefits. Consequently, how to select cancer patients to carry out costly immunotherapy is our responsibility and also an art to be practiced. ⁹⁰