Clinical diagnosis and treatment of immune checkpoint inhibitor‐associated adverse events in the digestive system

Immunotherapy for malignant tumors is a hot spot in current research and the treatment of cancer. The activation of programmed cell death receptor‐1 (PD‐1) and cytotoxic T lymphocyte‐associated antigen 4 (CTLA)‐4 relevant signaling pathway can inhibit the activation of T lymphocytes. Tumor cells can achieve immune escape by activating this signaling pathway. By inhibiting this signaling pathway, immune‐checkpoint inhibitors (ICIs) activate T lymphocytes to clear the tumor cells. Therefore, the adverse effects of ICIs are mainly immune‐related adverse events (irAEs). The digestive system, including the gastrointestinal tract and liver which are vital organs of digestion and absorption, metabolism and detoxification, as well as important immune‐related organs, is the most commonly affected system of irAEs. This review explains the incidence, clinical features, diagnosis and treatment of liver and gastrointestinal adverse events in ICIs.


Introduction
Checkpoint inhibitors (ICIs) can be classified into three categories based on the mechanism: (i) CTLA-4 inibitor: for example ipilimumab, which is a monoclonal antibody targeting CTLA-4; (ii) PD-1 inhibitors: pembrolizumab and nivolumab are anti-PD1 monoclonal antibodies; (iii) PD ligand 1 (PD-L1) inhibitor: atezolizumab, avelumab and durvalumab are monoclonal antibodies which target PD-L1. To date, the efficacy of checkpoint inhibitors have been shown in treating a variety of latestage malignancies including metastatic melanoma, lung cancer, renal cell cancer, hematologic malignancies and so on. ICI toxicity may cause a wide range of immune-related adverse events (iRAEs) in any system. In this article, we review the toxicity of ICIs in the liver and gastrointestinal tract.

Immune-related hepatotoxicity Incidence
The incidence of immune-related hepatotoxicity varies between different ICIs, mono-or combined therapy, and the underlying malignancy. Hepatotoxicity occurs in 5%-30% of patients receiving ICI therapy. Hepatotoxicity has been reported in up to 15% of patients receiving CTLA-4 inhibitors, while in 5%-10% of patients with PD-1/ PDL1. 1 Hepatitis occurred in 25%-30% (of which 15% was grade 3) of patients treated with combination therapy (ipilimumab combined with nivolumab). 2,3 The onset of hepatotoxicity has been reported to occur at any time after therapy but is most commonly seen after 8-12 weeks of treatment. 4

Diagnosis
The onset of hepatotoxicity in irAE is usually insidious without obvious clinical manifestation. Routinely monitoring the liver function tests before and after every cycle of treatment is crucial for early detection. The diagnosis of ICI-related hepatotoxicy can be very challenging, since there is neither specific clinical features nor diagnostic biomarkers, and other etiologies can be coexisting such as liver malignant metastasis, infections and hepatotoxicity caused by concurrent medications.
Before ICI therapy It is necessary to document any history of chronic liver disease including drinking habits, long-term history of hepatotoxic medicine use, history of chronic hepatitis and autoimmune liver disease before treatment initiation. Antibodies related to autoimmune liver disease and tests for viral hepatitis including hepatitis B and hepatitis C viruses should be screened before commencing treatment.
During and after ICIs therapy Once abnormal liver function tests have been reported, or increased compared with baseline level, a series of tests including blood biochemistry, tests for viral hepatitis, liver imaging examination and liver biopsy when needed should be completed as soon as possible. An increase in ALT or AST of more than two times the upper limit of normal should prompt workup of hepatotoxicity. Serological markers of hepatitis virus, including HAV, HBV, HCV, and opportunistic infection pathogens (CMV and EBV) should be established. Additional workup should include antinuclear antibodies (ANA), and smooth muscle antibody (SMA) tests. Medication reconciliation including evaluation for alternative therapy/herbal medication should be performed and any potential hepatotoxic medications discontinued.
Abdominal imaging Abdominal imaging including computed tomography (CT), magnetic resonance imaging (MRI), and ultrasound (US) are usually nonspecific for the diagnosis of ICI-related hepatitis. However, imaging evaluation can be of value in the diagnosis of metastatic liver disease and thromboembolic events. Reported imaging features of ICI-related hepatitis include hepatomegaly, periportal edema, periportal MRI T2-hyperintensity, attenuated liver parenchyma, and enlarged periportal lymph nodes on CT and MRI in severe hepatitis. 5 In patients presenting with cholestasis (increased bilirubin, mainly conjugated bilirubin, and increased GGT and ALP), bile duct obstructive factors including biliary stone, tumor, or periportal lymphadenopathy should be excluded by abdominal imaging examination (ultrasound, MRCP, etc). To ensure that the diagnosis is confirmed in a timely manner and avoid delaying therapy for hepatotoxicity, identification of the above causes should be initiated early, and comprehensive evaluation before initiating ICIs is a priority.
Liver biopsy A liver biopsy should be considered in patients with grade 4 adverse events (Table 1) to assist with a differential diagnosis and predicting prognosis. Histological examinations of ICI-related hepatitis include nonspecific features of panlobular hepatitis indistinguishable from autoimmune hepatitis, bile duct injury including fibrin ring granulomas, central vein endotheliitis, prominent sinusoidal lymphohistiocytic infiltrates, and endotheliitis involving the central vein. 6,7 It has been reported that CTLA-4 inhibitor-induced hepatitis caused granulomatous hepatitis with fibrin deposits, whereas PD-1/PD-L1 inhibitor induced hepatitis caused lobular nongranulmatous hepatitis. 4 A recent study has also revealed that ICI-induced hepatitis showed increased numbers of CD3+ and CD8+ lymphocytes and decreased CD20+ B cells and CD4+ T cells compared with autoimmune hepatitis and drug-induced liver injury. 8

Management
The management principles of hepatotoxicity are shown in Table 1, based on the severity according to elevated level of transaminase (ALT, AST) and whether total bilirubin, INR, or albumin abnormalities were present. ICIs can usually be continued in patients confirmed to have a grade 1 hepatotoxicity. In patients with grade 2 hepatotoxicity, consideration should be given to withholding ICIs. In patients with grade 3 or Grade 4, permanent withdrawal of ICIs should be considered. Corticosteroids are the preferred therapy for treating hepatotoxicity. In patients with grade 3 or grade 4 hepatitis whose liver function tests decreased to grade 2 after IV methylprednisolone, treatment can be downgraded to oral prednisolone, and then gradually tapered within four weeks thereafter. In patients with grade 2 hepatitis who respond well to oral prednisolone, corticosteroids can be tapered within two weeks. In patients with progressively aggravated liver function during corticosteroid treatment, intensive upgraded treatment should be considered. When there is no response to intravenous methylprednisolone 2 mg/kg/day in two to three days, oral mycophenolic acid (MMF) 500-1000 mg bid should be added. Tacrolimus may be considered in case of failure after the administration of MMF. It has also been reported that antithymocyte globulin (ATG) can be used to treat severe hepatitis in patients who have failed to respond to corticosteroids. 9 ICI-related hepatotoxicity usually resolves within four to six weeks with the appropriate treatment. In the event that it does not resolve, it is necessary to be alerted to other contributory factors and the initial diagnostic workup should be repeated, bearing in mind the concurrent medications with potential hepatotoxicity, reactivation of CMV, as well as other causes.

Gastrointestinal toxicity of ICIs Incidence
The gastrointestinal (GI) tract is the most commonly affected system with irAEs compared with other organs or  systems. The incidence of GI irAE differs between different targets of ICIs. GI events caused by CTLA-4 inhibitors are higher than that of PD-1/PD-L1 inhibitors. The incidence of GI irAEs also differs in patients with tumors of different origin. 10 The incidence of GI irAEs has been reported to be higher in melanoma patients treated with PD-1 inhibitors than that of patients with non-small cell lung cancer (NSCLC) and renal cell cancer. It suggests that the same drug may drive the histologically specific irAE pattern in different immune microenvironments. 10 It has been reported that diarrhea occurred in 27%-54% of cancer patients treated with CTLA-4 inhibitors, and colitis in 8%-22%. 11 GI adverse events are the most commonly reported irAEs of CTLA-4 inhibitors, with the earliest onset and very severe symptoms which often leads to treatment discontinuation. It has been reported that 1%-1.5% of melanoma patients treated with ipilimumab developed colonic perforation, and that of renal cell carcinoma patients was 6.6%. 12 The incidence of grade 3/4 GI irAEs of PD-1 inhibitors was 1%-2%. 2 A GI irAE may appear at any time from the first to the tenth dose of CTLA-4 inhibitor, even several months after it was last administered. 13 The median onset time of GI irAEs has been reported to be one month after initiation of CTLA-4 inhibitor, and 2-4 months after PD-1 inhibitor. 14 GI irAEs occur more frequently, severely and early in patients with a combination treatment of CTLA-4 inhibitors and PD-1 inhibitors. 2 The risk factors of GI irAEs include use of NSAIDs and medical history of inflammatory bowel disease. 13,15 Diagnosis Diarrhea is the most common manifestation of GI irAEs. Other symptoms include abdominal pain, blood in the stool, nausea, vomiting, weight loss, and fever. It can be accompanied by a variety of extraintestinal manifestations, such as arthralgia, endocrine disorders, skin disorders, hepatitis, nephritis, pericarditis, pancreatitis and other irAEs. Laboratory examination may show elevated levels of C-reactive protein, anemia, and hypoalbuminemia. Positive autoimmune antibodies such as antineutrophil cytoplasmic antibody (ANCA) etc have been detected in a minority of patients. 13 Endoscopic features are mucosal erythema, loss of vascular pattern, erosions and ulcerations. The left-side of the colon is involved in most cases. The lesions can be diffuse or discontinuous. Histopathological features usually present as acute injury pictures (infiltration with neutrophils and eosinophils), either focal or diffuse, with crypt abscesses. In some cases, the histopathological features of chronic inflammation including disruption of crypt structures (branching, atrophy, budding, distortion, etc), basal plasmocytosis and even granuloma has been observed. 13 There are similarities and overlaps in clinical, endoscopic and even pathological features between GI irAEs and inflammatory bowel disease (IBD), a chronic nonspecific digestive disease. Table 2 summarizes the abovementioned features and the differences compared with IBD. 16 The diagnosis of GI irAEs depends on the time relationship between the development of clinical symptoms and the administration of ICIs in patients with the above laboratory, endoscopic and histopathological features. Other causes should simultaneously be excluded, including infectious enteritis, such as Clostridium difficile infection, CMV colitis and other opportunistic infections; ischemic colitis; colitis caused by other medication such as NSAIDs; radiation enteritis and so on. Therefore, it is recommended to perform a stool pathogen examination including stool routine test, Clostridium difficile toxin, stool microscopy for leukocytes/parasites/ova, culture for drug-resistant organisms, and viral PCR (serum CMV-DNA). Meanwhile, an enhanced CT scan of the abdomen and pelvis, as well as sigmoidoscopy or colonoscopy with biopsy are recommended based on consultation with a gastroenterologist.

Management
The principles of managing GI irAEs are early identification, timely and adequate treatment, and rapid escalation to improve prognosis. Treatment is stratified on the severity of diarrhea which is graded according to the frequency of bowel movements. Table 3 shows the treatment strategies of GI irAEs based on diarrhea severity. Corticosteroids are the main therapy for moderate and severe GI irAEs. In patients with a full response to corticosteroids, treatment can usually can be tapered within two to four weeks in patients with moderate GI irAEs and four to eight weeks in patients with severe symptoms. In patients with inadequate or no response to corticosteroids, dose escalation of steroids in time, and upgrading treatment to infliximab (IFX) or vedolizumab when necessary is recommended. Studies have shown that compared with long-term steroid treatment, the therapy of short-term steroids plus IFX reduces the risk of various opportunistic infections. 17 Fecal microbiota transplant is reported to be valid in cases with GI irAEs refractory to steroids, IFX or vedolizumab. 18 Intestinal microbiota, GI irAEs and tumor prognosis Biomarkers that predict gastrointestinal irAEs have so far not been identified. Based on the latest research, fecal microbiota composition at baseline before treatment with ICIs predicts ipilimumab-induced colitis. Studies have shown that the intestinal microbiota enriched with clostridium and other firmicutes at baseline is related to a good response to ipilimumab and higher incidence of immunerelated colitis. 19 Several studies published in early 2018 reported that some specific fecal microbiota features prior to treatment have been associated with a good response to ICIs, which bring further prospects for enhancing the efficacy of ICIs in treating patients with cancer and improving prognosis. 20,21

Conclusions
With the widespread use of ICIs in cancer therapy, irAEs are increasingly being valued by oncologists and specialists.
The gastrointestinal system (diarrhea, colitis) is usually most affected, followed by liver involvement in digestive system irAEs. Baseline screening, early identification, timely diagnosis, rapid and adequate treatment of irAEs are key to addressing this type of clinical problem.
Whether the composition of intestinal microbiota can predict GI irAEs and predict the prognosis of ICIs in treating tumors deserves further research and exploration.