Monoclonal Antibodies in Cancer

Abstract

The development of hybridoma technology by Kohler and Milstein 1975 was a milestone in the development of monoclonal antibody technology. It was possible now, to generate unique, uniform monoclonal antibodies (MABs) with a defined specificity and a reproducible quality. In 1979 the first patient was treated with a therapeutic MAB, which had until then only been used as diagnostic tools. In this chapter, the different classes and characteristics of MABs used as cancer therapeutics are described. Safety aspects of selected antibodies are discussed as well.

Introduction

More than 60 years ago the first chemotherapeutic drugs were used to support surgery and/or radiation therapy against cancer. In the following decades, chemotherapy developed to a mainstay of therapy rather than being supporting in character. In the last two decades monoclonal antibodies (MABs) have faced a growing interest as antitumor therapeutics. In November 1997, Rituximab was the first-in-class therapeutic MAB approved by the FDA. In June 1998 followed the approval in the European Union. At that time, even the manufacturer believed the approval as monotherapy for relapsed/refractory, CD20 positive, low-grade or follicular B-cell Non-Hodgkin’s lymphoma would be a niche indication. The following rapid development of many different MABs in multiple different cancer indications shows how incorrect this assessment was. Nowadays, a large number of MABs has been approved for use in numerous cancer indications with many more in different stages of clinical development.

Classification of Monoclonal Antibodies

Structure, functionality, classes and possible modifications of MABs and their mode of action are described in Chap. 8. MABs can be roughly divided into unconjugated antibodies and conjugated antibodies . The latter ones can be subdivided into toxin coupled antibodies—so called chemoimmunoconjugates or antibody drug conjugates (ADC) —and radionuclide coupled antibodies , the radio immune conjugates (RIC). These “naked”, uncoupled toxins given alone would be too toxic for the patient. The antibody works as a tugboat, tugging the toxin to accumulate selectively nearby or in the tumor. A RIC can be considered “inner radiation therapy”.

Pharmacological Targets of Oncologic Antibodies

MABs can be directed against surface structures of malignant cells. These structures comprise receptors or proteins, such as the epidermal growth factor receptor (EGFR) or the cluster of differentiation (CD) antigens. Besides that, some MABs intercept soluble (growth) factors in the blood and peripheral tissue fluids, which could lead to a tumor survival benefit. One example in oncology is the vascular endothelial growth factor (VEGF), which is intercepted by bevacizumab or aflibercept (“VEGF-Trap”). The most recent and rapidly growing class of MABs are immune agonistic antibodies, currently directed against the cytotoxic T-lymphocyte antigen 4 (CTLA4), the programmed cell death receptor1 (PD 1) or against its ligand (PD-L1). The main difference in the mode of action compared to all the other classes of MABs and even chemotherapeutics is that not the tumor itself is attacked, but the immune system of the patient is held “under fire” to re-recognize the tumor. Table 23.1 gives an overview over the present MABs with an oncology indication, ordered by the target structure.

Table 23.1 Therapeutic MABs clustered by their targets

All mentioned targets can also be found on non-malignant tissues, which explains the observed toxicity. At present time, none of the approved MABs is directed against a tumor specific molecular driver, as they are known for instance in Philadelphia chromosome positive CML (the fusion protein BCR/Abl), in anaplastic lymphoma kinase (Alk) positive non-small cell lung cancer (NSCLC), in EGFR mutated NSCLC (mutated EGFR), or in the FLT3-positive AML. Her2 (=ErbB2), for example, can also be found in normal tissues, but in Her2 positive breast cancers it is overexpressed.

Pharmacokinetics of Mabs: General Remarks

Unlike small, defined molecules , the pharmacokinetic properties of MABs differ markedly. Distribution into tissue is slow (internalization) because of the molecular size of MABs. Volumes of distribution are generally low. Metabolism is similar to catabolic degradation of endogenous and dietetic peptides and proteins. The half-life is usually relatively long, which allows for long dosing intervals during maintenance therapy. Rituximab, for instance, is given every 2 months in the first line setting and every 3 months in the relapsed or refractory situation. Possible factors influencing the pharmacokinetics include the amount of the target antigen, which corresponds to the total tumor mass, immune reactions to the antibody (anti-drug antibodies) and patient demographics such as gender or age (see section on rituximab). Population pharmacokinetic analyses have been applied in assessing covariates in the disposition of MABs. Both linear and nonlinear elimination have been reported for MABs, which is probably caused by target mediated disposition (overview Keizer et al. 2010). If a tumor responds to a MAB therapy, the amount of the remaining target antigen diminishes and the half-life can be prolonged. However, MAB dosing is based on different pharmacokinetic models, leading to approved dosing strategies of body surface area-based (rituximab), body weight-based (trastuzumab, pertuzumab) or flat dosing (atezolizumab, obinutuzumab ).

Anti-CD Antibodies

The cluster of differentiation (also known as cluster of designation or classification determinant and often abbreviated as CD) denotes groups of immune phenotyping surface properties for the identification and investigation of cell surface molecules providing targets on cells in order to classify these cells according to their biochemical or functional characteristics. CD molecules are membrane bound proteins, often glycosylated, in part cell specific. In terms of physiology, CD molecules can act in numerous ways, often acting as receptors important to cell functioning and/or survival. Others have enzymatic activity. A signaling cascade is usually initiated or modulated, altering the behavior of the cell. Additionally to the mechanisms of ADCC and CDC (Chap. 8), MABs can alter or even block signal transduction pathways, leading to cell death via apoptosis (overview Ludwig et al. 2003).

■ Anti CD20 Antibodies

In the 1980s, CD20 was identified as a B-cell marker by Stashenko et al. (1980). CD20 is expressed on early pre-B cells, it remains through B-cell development, and is then lost from plasma cells (Tedder and Engel 1994). It is not found on hematopoietic stem cells. Therefore, anti CD20 antibodies are also designated as anti-lymphocytic antibodies. The antigen is neither shed nor internalized. CD20 is expressed on the majority of B-cell lymphomas (Stashenko et al. 1980; Tedder and Engel 1994; Nadler et al. 1981). It is a nonglycosylated member of the membrane-spanning 4-A (MS4A) family (Ishibashi et al. 2001). The antigen consists of three hydrophobic regions forming a tetraspan transmembrane molecule with a single extracellular loop and intracellular N- and C-terminal regions (Tedder et al. 1988). CD20 has been shown to be present on the cell surface as a homo-multimer, in tetramer complexes (Polyak and Deans 2002; Polyak et al. 2008). Despite decades of research, no natural ligand for CD20 could be detected (Cragg et al. 2005). However, subsequent data revealed that CD20 is resident in lipid raft domains of the plasma membrane where it probably functions as a store-operated calcium (SOC) channel following ligation of the B cell receptor (BCR) with the antigen, implying that it plays a role in regulating cytoplasmic calcium levels after antigen engagement (Bubien et al. 1993; Kanzaki et al. 1995; Li et al. 2003).

■ Pharmacology

Anti-CD20 antibodies can be classified into type I and type II antibodies with different pharmacodynamics (Cragg and Glennie 2004). Type I (rituximab-like) MABs induce CD20 to redistribute into large detergent resistant microdomains (lipid rafts), whereas type II (tositumomab-like) MABs do not (Deans et al. 1998). These lipid microenvironments on the cell surface—known as lipid rafts —also take part in the process of signal transduction. Lipid rafts containing a given set of proteins can change their size and composition in response to intra- or extracellular stimuli. This favors specific protein–protein interactions, resulting in the activation of signaling cascades. For details see Simons and Toomre (2000). Rituximab and ofatumumab are type I, whereas obinutuzumab is the first glyco-engineered type II MAB. Type I antibodies display a remarkable ability to activate complement and elicit complement-dependent cytotoxicity (CDC, Chap. 8). This is a result of enhanced recruitment of the complement factor C1q. This ability, however, appears to be directly linked to their (CD20 bound) translocation into lipid rafts, which cluster the antibody Fc regions thus enabling improved C1q binding (Cragg et al. 2003). Type II MABs do not show these characteristics: They do not change CD20 distribution after binding; no concomitant clustering is observed and they are relatively ineffective in CDC. Interestingly, they evoke far more homotypic adhesion and direct killing of target cells in a caspase independent manner (Chan et al. 2003; Ivanov et al. 2008). In summary, type I MABs have to cross-link the homo-tetrameric CD20 antigen for a pharmacodynamic effect. In diseases like CLL, with a lower density of CD20 antigens compared with other B-cell lymphomas, the tetramers have to be bridged with more than one molecule of rituximab. That is the explanation why rituximab in CLL is given at a higher dose of 500 mg/m² in subsequent cycles after the usual starting dose of 375 mg/m². For illustration see Fig. 23.1.

Figure 23.1

Anti-CD20 MABs can be divided into two distinct subtypes, termed type I and II. Type I MABs induce CD20 to redistribute into large detergent resistant microdomains (rafts) whereas type II MABs do not. This differential ability of anti-CD20 mAbs to redistribute CD20 in the cell membrane impacts on many of the binding properties and effector functions that control the therapeutic success of anti-CD20 mAbs. Type I and II MABs have the ability to evoke different effects: type I MABs engage ADCC and cause CD20 modulation but do not elicit direct cell death, whereas type II MABs mediate direct cell death and engage ADCC, but do not promote CD20 modulation (Beers et al. 2010). In diseases with a low CD20 density, more than one rituximab molecule has to bridge the CD20 antigens

Type II MABs bind within a CD20 tetramer, without lipid raft redistribution. They induce a “closed conformation” (Beers et al. 2010; Niederfellner et al. 2011) (Fig. 23.2). The antigens don’t have to be bridged. The type II MAB-induced cell death is dependent on homotypic adhesion, requires cholesterol, and is energy-dependent, involving the relocalization of mitochondria to the vicinity of the cell–ell contact (Beers et al. 2010).

Figure 23.2

Typ II MAB (Tositumomab like) bind within a CD20 tetramer, and the antigens don’t have to be bridged. No redistribution into lipid rafts occurs (Beers et al. 2010; Niederfellner et al. 2011)

The pharmacodynamic mechanism of action and the resulting effects of rituximab are illustrated in Fig. 23.3 (from Stolz and Schuler 2009).

Figure 23.3

Overview of rituximab-inducible indirect (ADCC und CDC) effector mechanisms and intracellular, CD20 triggered signal transduction cascades after CD20 cross linking (Stolz and Schuler 2009)

Rituximab

Rituximab is used as a monotherapy as well as in combinations with chemotherapy. It has become a mainstay in the treatment of B-cell malignancies such as aggressive and indolent non-Hodgkin’s lymphomas (NHLs) and chronic lymphocytic leukemia (CLL), as well as other B-cell triggered diseases. It is also widely utilized in an off label fashion for numerous clinical conditions like immune thrombocytopenia (ITP). Histology subtype has been described as the main tumor parameter that influences rituximab efficacy. In an early trial, when rituximab was given for four doses as monotherapy, small lymphocytic lymphoma histology negatively influenced the objective response rate compared with other low-grade lymphomas (McLaughlin et al. 1998). Several studies have described a low objective response rate in patients with CLL treated with the standard dose of 375 mg/m² rituximab monotherapy used in other NHL (Huhn et al. 2001; Nguyen et al. 1999; O’Brien et al. 2001). As mentioned above, the level of CD20 expression (density) may differ depending on tumor histology (Manches et al. 2003). This was initially used to explain the variation in rituximab efficacy among different histological subtypes. Despite conflicting in vitro data, the dosing in CLL was adjusted to 500 mg/m² for subsequent doses after an initial dosing of the well-known 375 mg/m² finally. The reduced dose for the first infusion is necessary because of an usually high tumor load in need for therapy in CLL blast crisis. Responding to rituximab therapy bears a high risk of tumor lysis and cytokine release syndrome as well as other infusion related reactions (IRR); see also under ‘safety’. For induction therapy as well as for maintenance therapy (used in follicular lymphoma—FL) a minimum serum concentration has to be achieved for tumor response (Berinstein et al. 1998; Gordan et al. 2005).

Responses to therapeutic MABs have also been reported to correlate with specific polymorphisms in the Fcγ receptor (FcγR), especially when ADCC is a major part in the mechanism of action (Mellor et al. 2013). These polymorphisms are associated with differential affinity of the receptors to MABs. At present, these functional FcγR polymorphisms are not suitable to be used as pharmacogenetic biomarkers that could be used to better target the use of MABs in cancer patients. The available studies do not describe a consistent effect of FcγR genotype on the clinical anti-tumor activity of therapeutic IgG1 MABs. Inconsistencies observed in studies are likely related to differences in tumor type, the cytotoxic agents used in combination, the clinical settings, like metastatic vs. adjuvant situation, the clinical benefit parameters measured, the therapeutic antibody used, as well as the magnitude and even the direction of the effect. However, even patients with an “unfavorable” FcγR genotype do benefit from an antibody containing regimen (overview Mellor et al. 2013).

To minimize IRR, the infusion rate is increased slowly from the initial infusion rate of 50 mg/h every 30 min by doubling the subsequent infusion rate if no reactions occur up to 400 mg/h. Shortly before approval of rituximab biosimilars, a fixed dose, subcutaneous (s.c.) formulation was launched in the EU. Rituximab is concentrated up to 120 mg/mL in recombinant hyaluronidase, a penetration enhancer (“spreading factor”). The dose for follicular lymphoma is 1400 mg (12 mL), for CLL 1600 mg (16 mL). The first dose has still to be given by the i.v. route due to tolerability concerns. The subsequent s.c. doses can be given over 6 min—an substantial advantage from the patient’s point of view.

As shown by the SABRINA study, the pharmacokinetic profile of s.c. rituximab was non-inferior to i.v. rituximab. I.v. and s.c. rituximab had similar efficacy and safety profiles, and no new safety concerns were noted. S.c. administration does not compromise the anti-lymphoma activity of rituximab when given with chemotherapy.

Rituximab Safety

From today’s point of view, dose finding and scheduling of rituximab administration, the first approved therapeutic anti-cancer antibody, were more empirical than rational or biologic target based. As no dose-limiting toxicity or clear dose–response relationship was found during a phase I study with single doses of 10, 50, 100, 250 or 500 mg/m² (Maloney et al. 1994), the 375 mg/m² dose level with a 4 weekly dosing scheme was chosen for phase II studies (McLaughlin et al. 1998; Maloney et al. 1997) without an apparent reasoning. A dose escalation trial was conducted in CLL and even at 2250 mg/m² (monotherapy) the maximum tolerated dose was not reached (O’Brien et al. 2001). Rituximab has significant activity in patients with CLL at the higher dose levels. However, the dose-response-curve is not steep. Myelosuppression and infections are uncommon.

Rituximab is generally well tolerated, in patients with both malignant and non-malignant disease, including children (Quartier et al. 2001) and even pregnant women (Herold et al. 2001). Severe adverse events are rare but possible. The most common adverse events are infusion-/foreign protein-related and occur most frequently during or shortly after the first infusion. However, extremely rare, hypersensitivity reactions can occur even after years of maintenance therapy, leading to permanent discontinuation of its use. This syndrome consists of chills, fever, headache, rhinitis, pruritus, vasodilation, asthenia and angioedema. A cytokine release syndrome is possible. Less often reported are hypotension, rash, bronchospasm, pain at tumor sites and rash or severe skin toxicity inclusive Stevens-Johnson-Syndrome and Toxic Epidermal Necrolysis (=Lyell-Syndrome). Routine premedication consists of corticosteroids, if they are not already a component of the chemotherapy- or antiemetic scheme. The same drug classes are used for intervention in case of hypersensitivity reactions (HSR) despite correct premedication.

Rituximab induces a rapid depletion of CD20⁺ B-cells in the peripheral blood. B-cells remain at low levels for at least 2–6 months with recovery to pretreatment values occurs within 12 months (Maloney et al. 1997). Other hematological toxicities comprise temporary reduction of platelets or neutrophils, and occasionally reduced immunoglobulin levels, which also can be caused by the underlying disease and its bone marrow involvement. Despite the fact that the drug is generally well tolerated, a small number of patients experience unexpected and severe toxicities. It is speculated, that these patients have a rituximab serum level dramatically above the average. However, so far, this could not be proven in clinical routine.

Also, gender and age seem to influence rituximab kinetics and clinical outcome. The German High-Grade Non-Hodgkin Lymphoma Study Group (DSHNHL) first detected in elderly women with diffuse large B cell lymphoma (DLBCL) a slower elimination of rituximab compared with men, which translated in a better outcome for these women (Pfreundschuh et al. 2014). Consequently, the DSHNHL conducted a phase II trial increasing the number of rituximab infusions to achieve high rituximab levels early during treatment. In this DENSE-R-CHOP-14 trial, 100 elderly patients with aggressive CD20⁺ B-cell lymphoma received 6 cycles of biweekly CHOP-14 combined with 12 x rituximab (375 mg/m²) on days 0, 1, 4, 8, 15, 22, 29, 43, 57, 71, 85, and 99. (CHOP-14 treatment regimen: Cyclophosphamide 600 mg/m² day 1; doxorubicin (=Hydroxydaunorubicin) 50 mg/m² day 1; Vincristine (Oncovin®) 1.4 mg/m² max. 2 mg day 1; Predniso(lo)ne 100 mg day 1–5 every 14 days).

This intensification of rituximab administration achieved higher rituximab serum levels and resulted in higher complete remission and event-free survival rates in elderly patients with poor-prognosis DLBCL (Pfreundschuh et al. 2007). The negative impact of male gender in DLBCL was confirmed by a meta-analysis (Yildirim et al. 2015). While body weight contributes to a faster elimination in males according to (Muller et al. 2012), others found a significant longer five-year OS in a high BMI group (>22.55 kg/m²) when compared to that of the low BMI group (Weiss et al. 2017).

With the increasing use of Rituximab, a growing number of rare adverse effects have been recognized. Late-onset neutropenia (LON) is, according to the National Cancer Institute Common Toxicity Criteria defined as grade III–IV neutropenia, in which absolute neutrophil count is less than 1.0 × 10³/L, occurring 4 weeks after the last rituximab administration. LON has been reported in 5–27% of rituximab-treated lymphoma patients. Similar figures apply for autoimmune patients but those appear to have more infections during the neutropenic period (Tesfa et al. 2011; Tesfa and Palmblad 2011).

The mechanisms of LON after rituximab treatment still has not been elucidated. One hypothesis is, that LON may result from hematopoietic lineage competition due to an excessive B-cell recovery in the bone marrow by promotion of B-cell lymphopoiesis over granulopoiesis (Anolik et al. 2003). Others see a correlation between a high-affinity FCGR3 158 V allele and LON in lymphoma patients (Weng et al. 2010). A clinically relevant question is if it is safe to re-treat patients with rituximab who previously developed LON. Recurrence of LON episodes upon re-exposed patients has been reported. Nevertheless, a close clinical follow-up or complete blood count monitoring is considered adequate in most LON cases (Tesfa and Palmblad 2011).

Following Rituximab administration, reactivation of hepatitis B, in some cases resulting in fulminant hepatitis, hepatic failure, and death, has been reported. This led to testing of active replicating hepatitis as well as to screening of all patients for HBV infection by measuring HBsAg and anti-HBc before initiating treatment with rituximab as a therapeutic precondition. Monitoring and/or reactivation of antiviral prophylaxis should be considered, and is advised by the FDA’s Full Prescribing Information (FPI) and the European Medicines Agency’s Summary of Product Characteristics (SmPC). HBV reactivation can occur up to 24 months following completion of rituximab therapy.

The immune suppressing antibody ± chemotherapy can lead to reactivation of latent JC polyoma virus, leading to potentially fatal Progressive Multifocal Leukoencephalopathy (PML) . Patients presenting with onset of neurologic manifestations should consult a neurologist with this suspected diagnosis. For natalizumab, an anti-alpha4-integrin antibody used against multiple sclerosis, that induced PML in some patients, removal of the drug via plasma exchange was an important part of the therapy, as it reconstituted the immune system. In a case report, the first successful removal of rituximab with plasma exchange therapy (rituximab apheresis) and accompanying complement-dependent cytotoxicity test (CDC) assay for monitoring, has been described. Neurologic symptoms of the patient improved within the first 2 weeks (Burchardt et al. 2012).

Ofatumumab

Ofatumumab is a type I, human immunoglobulin (Ig) G1k antibody. Ofatumumab binds with greater avidity than rituximab to another epitope of CD20, which encompasses the small extracellular loop (residues 74–80) and the N-terminal region of the second large extracellular loop (Fig. 23.4). Several crystal structure based analyses of the Fab fragment of the antibody suggests that ofatumumab can bind closer and tighter to the cell membrane than rituximab, leading to more effective CDC. Induction of CDC appears to be greater than with rituximab and occurs even at a lower density of CD20 on the cell surface (Cheson 2010). After binding to CD20 on malignant B cells, ofatumumab induces clustering of CD20 into lipid rafts, similar to other type I antibodies as described above. In contrast to rituximab, ofatumumab does not induce apoptosis of B-cell lines (Cheson 2010).

Figure 23.4

Distinct CD20 binding epitopes of rituximab, ofatumumab and obinutuzumab (designated as GA101). Ofatumumab binds to the so called “small loop” (from Klein et al. 2013)

Initially licensed for double refractory CLL, namely those resistant or refractory to fludarabine and alemtuzumab, ofatumumab can be used in previously untreated as well as relapsed patients after conventional chemotherapy. As already mentioned, CLL in need for therapy (blast crisis) bears a risk of rapid cell breakdown in the sense of a tumor lysis syndrome. Therefore a reduced initial flat dose of 300 mg is given. The subsequent flat dose is 1000 mg or 2000 mg, depending on the clinical situation (details see FPI and SmPC).

Ofatumumab Safety

Qualitatively the toxicity profile of ofatumumab is similar to that of rituximab. This includes HBV reactivation and the potential for PML. No additional toxicity or safety signals have been observed so far.

Obinutuzumab

Obinutuzumab is the first glycoengineered, type II humanized anti-CD20 MAB. It is post-translational defucosylated, resulting in the absence of a fucose sugar residue from IgG oligosaccharides in the Fc region of the MAB molecule (Tobinai et al. 2017). Obinotuzumab binds within the CD20 tetramer, as depicted in Fig. 23.2. CDC, probably, does not contribute to the overall activity of obinutuzumab. The limited capacity of obinutuzumab to fix complement via its Fc portion may further enhance its capability to bind to FcγRIII and mediate ADCC (summarized in Tobinai et al. 2017). Obinutuzumab was significantly more effective than rituximab in depleting B cells in whole blood samples from healthy donors (n = 10) and from an individual with CLL (Mossner et al. 2010). In CLL, the dosing of obinutuzumab is 100 mg on day 1, followed by 900 mg on day 2 in cycle 1. If tolerated and no IRR occur, the second part of the split dose can be given on day 1. In subsequent cycles there is no dose splitting on day 1. As mentioned before, starting therapy of CLL may lead to the tumor lysis syndrome. Therefore, the split dose is a safety measure and not necessary in other diseases, e.g. indolent NHL. According to the FPI and SmPC no dose splitting is advised for follicular lymphoma.

In contrast to rituximab, obinutuzumab is ramped up in cycle 1 with a flat dose of 1000 mg on days 1, 8 and 15. Consequently, patients in head-to-head comparisons of obinutuzumab with rituximab received 3000 mg obinutuzumab in cycle 1, whereas rituximab patients received 375 mg/m². This was often seen as a dosing imbalance. However, the manufacturer did not study a second experimental arm with the same flat dose for rituximab, which would have been considered a real head-to-head study. Different dosing regimens were tested for obinutuzumab. For achieving the desired serum concentration, 1600 mg on days 1 and 8 in cycle 1, followed by 800 mg on day 1 in subsequent cycles was determined. 1600 mg might need very long infusion times due to possible IRR, especially in CLL. Results in the GAUGUIN study (Morschhauser et al. 2013) and pharmacokinetic modeling showed that obinutuzumab 1000 mg per cycle with additional 1000 mg doses on days 8 and 15 of cycle 1 can achieve exposures similar to the 1600/800 mg regimen. This simplified flat-dose 1000 mg schedule was adopted for subsequent phase II and III investigations (Cartron et al. 2016).

Obinutuzumab Safety

The safety profile of obinutuzumab is similar to the other anti-CD20 antibodies rituximab and ofatumumab.

Y⁹⁰ Ibritumumab and I¹³¹ Tositumumab Tiuxetan

Y ⁹⁰ Ibritumumab and I ¹³¹ Tositumumab Tiuxetan are murine RICs. Yttrium-90 is a beta-emitter; Iodine-131 is a beta-emitter and an emitter of gamma radiation. Their activity is mainly achieved through their radioisotopes rather than their intrinsic antibody activity (Fig. 23.5). The radiation can penetrate through several cell layers, which is termed “cross fire” or “bystander” effect (Fig. 23.6). Adjacent cells, not marked by the antibody are also effected. Although active, these antibodies never gained a broad acceptance in the medical community. As of February 2014, the production of tositumomab and I¹³¹ tositumomab has been discontinued by the manufacturer and is no longer available (National Cancer Institute 2014), Y⁹⁰ ibritumumab tiuxetan is still approved in the EU, but rarely used.

Figure 23.5

Comparison of conventional radiotherapy (left) with an external radiation source. Radiation from inside (right) by RICs

Figure 23.6

“Cross fire” or “bystander” effect of radiation, penetrating more cell layers than marked by the antibody

■ Other Anti CD Antibodies: Unconjugated

Alemtuzumab

Alemtuzumab was another anti-lymphocytic antibody, directed against the CD52 antigen. The CD52 antigen is mainly found on mature B- and T lymphocytes, but can also be found on monocytes/macrophages, NK cells, eosinophils and epithelial cells of the male reproductive tract. Alemtuzumab was used for the treatment of double refractory B-CLL (refractory to an alkylator plus fludarabine). This hematological approval was retracted in 2012 to favor the development as an immunosuppressant against multiple sclerosis, for which it is currently approved.

■ Anti Myeloma Antibodies

Elotuzumab

Elotuzumab is an immunostimulatory monoclonal antibody that recognizes SLAMF7 (Signaling Lymphocyte Activation Molecule Family Member 7; CD319), a glycoprotein highly expressed on myeloma and natural killer cells but not on normal tissues (Hsi et al. 2008). Elotuzumab causes myeloma cell death via a dual mechanism of action (Collins et al. 2013) (Fig. 23.7).

• 1. Direct activation: Binding to SLAMF7 directly activates natural killer cells (Collins et al. 2013), but not myeloma cells (Guo et al. 2015).

• 2. Tagging for recognition: Elotuzumab activates natural killer cells via CD16, enabling selective killing of myeloma cells via antibody-dependent cellular cytotoxicity (ADCC) with minimal effects on normal tissue (Collins et al. 2013).

Figure 23.7

Dual mechanism of action of elotuzumab (Collins et al. 2013)

Elotuzumab is not active as a monotherapy (Radhakrishnan et al. 2017), so it is combined with an immune modulating drug, usually lenalidomide, and dexamethasone. This combination lowers the killing threshold for myeloma cells. Further combinations, for example with proteasome inhibitors, are currently being tested.

Daratumumab

Daratumumab is directed against the CD38 antigen, found on T cells (precursors, activated), B cells (precursors, activated), myeloid cells (monocytes, macrophages, dendritic cells), NK cells, erythrocytes and platelets. CD38 has multiple functions. It acts as a receptor in close contact with the B cell receptor complex and CXCR4. In engagement with CD31 or hyaluronic acid, it activates NF-kappaB, ZAP-70, and ERK1/2 pathways. It also works as an ectoenzyme. CD38 interacts with NAD⁺ and NADP⁺, which are converted to cADPR, ADPR, and NAADP, all intracellular Ca²⁺ mobilizing agents (Malavasi et al. 2011).

Because of marked quantitative differences in expression levels of CD38 between normal cells and leukemic cells, combined with its role in cell signaling, suggests that CD38 is an attractive target for immunotherapy treatment of multiple myeloma (MM) (Malavasi et al. 2008). It is highly and uniformly expressed on myeloma cells (Lin et al. 2004; Santonocito et al. 2004), whereas only a relatively low expression was detected on normal lymphoid and myeloid cells and in some tissues of non-hematopoietic origin (Deaglio et al. 2001). In summary, daratumumab binding to CD38 elicits a signaling cascade and immune effector function engagement, leading to (de Weers et al. 2011)

• Complement-dependent cytotoxicity (CDC)

• Antibody-dependent cell-mediated cytotoxicity (ADCC)

• Antibody-dependent cell-mediated phagocytosis (ADCP), a rather new/unknown mode of action (Overdijk et al. 2015)

• Induction of apoptosis

• Modulation of cellular enzymatic activities associated with calcium mobilization and signaling

• Combination of these activities leads to elimination of plasma cells from bone marrow in MM patients

Daratumumab also kills myeloid-derived suppressor cells—Tregs or negatively regulating T cells. A graphical overview of Daratumumab’s mode of action is presented in Fig. 23.8.

Figure 23.8

Modes of action of daratumumab (de Weers et al. 2011; Overdijk et al. 2015). Antibody mediated phagocytosis is a rather new pharmacodynamic process. Details see text

Daratumumab and Elotuzumb Safety

Both antibodies need an intensive premedication. Despite correct pre-treatment with antipyretics, antihistamines and multiple doses of oral and i.v corticosteroids, IRR occur during daratumumab and elotuzumab administration, which makes a close patient-monitoring mandatory. Especially for daratumumab, IRR can occur hours after completing the infusion. A suddenly stuffy nose, cough, throat irritation, allergic rhinitis, hoarseness during the infusion can be regarded as early signs of an upcoming IRR. Due to necessary infusion interruptions caused by these IRR, a s. c. formulation as described for rituximab may resolve the problem. Dissolved in recombinant hyaluronidase, the fixed dose of 1800 mg daratumumab (90 mL) achieves the same trough level on day 1 of cycle 3, compared to the i.v. dosage form.

Because CD38 is also expressed on erythrocytes, the indirect Coombs test used prior to blood transfusions is false positive, up to 6 months after the last infusion of daratumumab. Blood typing at baseline is therefore highly recommended before the first daratumumab infusion. Meanwhile, daratumumab in blood samples can be destroyed by incubation with dithiothreitol (DTT) prior to the blood compatibility test. DTT cracks the structure stabilizing disulfide bridges of the antibody.

Both, elotuzumab and daratumumab also interfere with serum protein electrophoresis or immune fixation assays, leading to false positive results in patients with IgGκ myeloma protein, making the assessment of the initial response difficult.

■ Bispecific Antibodies

Blinatumumab

Blinatumumab is a first-in-class bispecific antibody, directed against CD3 on T cells and CD19 on B cells. It is also designated as a BiTE , a Bispecific T cell Enhancer . This antibody construct is made of two single chain variable fragments (scFv) of anti CD3/CD19, enabling a patient’s T cells to recognize malignant B cells (Fig. 23.9). CD3 is part of the T cell receptor. CD19, originally a B cell marker, is expressed on the majority of B cell malignancies in normal to high levels. By linking these two cell types, T cells are activated to exert cytotoxic activity on the target cell. The usual clinical use is the Philadelphia chromosome negative B (precursor) ALL.

Figure 23.9

Two fused scFv of anti CD3/antiCD19 antibodies result in the blinatumumab construct. One arm of blinatumomab binds to CD3, the other binds to CD19. This engages unstimulated T cells, which destroys the CD19⁺ cells

Blinatumumab has a molecular weight of 54.1 kDa, approximately one-third of the size of a traditional monoclonal antibody (Wu et al. 2015). It is eliminated rather quickly by the kidneys with a half-life of 1.2 h. Therefore, blinatumumab has to be continuously infused over 1 month.

Blinatumomab Safety

Blinatumumab can cause cytokine release as well as fulminant neurological toxicities even with fatal outcome. Some of them were caused by prescription, calculation or handling errors. When changing the infusion lines, for instance, the used lines must not be flushed. This would result in an erroneous bolus application with severe toxicities or death. To avoid all this and to make blinatumumab therapy safer, the manufacturer developed risk minimization information brochures for physicians, other health care professionals and patients. The preparation protocol is rather complicated and a high-precision infusion pump is necessary.

■ Other Anti CD Antibodies: Toxin Conjugated

Antibody drug conjugates (ADC) are chemically linked combinations of MABs and small molecule drugs with anti-tumor activity. As mentioned above, the unconjugated drug alone is too toxic for the patient. Therefore, common features of toxin-coupled antibodies are:

• The ADC has to be stable in circulation in vivo

• The antigen has to be predominantly tumor specific

• After binding to the tumor antigen, the ADC has to be internalized

• When internalized, the ADC has to be degradable inside the cell to release the lethal cargo.

The linker between the MAB and toxin can be distinguished into enzymatically cleavable linkers, such as the dipeptide linker used in brentuximab vedotin, or enzymatically uncleavable linkers, such as the thioether linker in ado-trastuzumab emtansine (trastuzumab emtansine in the EU, syn. TDM1). The linker itself contributes to the properties of the ADC including PK and ADME (overview Han and Zhao 2014).

Gemtuzumab Ozogamicin

Gemtuzumab Ozogamicin (GO) is an anti-CD33 MAB, coupled with a cytotoxic enediyne -antibiotic N-acetyl derivate from Micromonospora echinospora ssp. Calichenensis, termed calicheamicin γ₁ ^I (gamma-one-iodine, Fig. 23.10).

Figure 23.10

The complex chemical structure of N-acetyl-calicheamicin γ₁ ^I coupled to an anti CD33 MAB (symbolized in the upper left; © J. Barth)

The calicheamicin cleaves double stranded DNA via a radical mechanism. Like bleomycin, it belongs to the so-called free radical-based-DNA-cleaving-natural-products. Calicheamicin binds with sequence specificity to TCCT-, TCTC- and TTT in the minor grove of the DNA (minor groove binding agent).

Gemtuzumab ozogamicin originally received accelerated approval in May 2000 as a stand-alone treatment for older patients with CD33-positive AML who had experienced a relapse. It was voluntarily withdrawn from the market after subsequent confirmatory trials failed to verify clinical benefit and demonstrated safety concerns, including a high number of early deaths. Gemtuzumab ozogamicin was re-approved in September 2017 with a modified dosing regimen. This approval includes a lower recommended dose, a different schedule in combination with chemotherapy or on its own, and a new patient population (FDA 2017).

Gemtuzumab Ozogamicin Safety

Worrisome adverse events are end organ damages such as veno-occlusive disease or the sinusoidal obstruction syndrome, which can lead to fatalities. Other, in part severe, treatment-emergent adverse events are hemorrhage, mucositis, nausea, vomiting, constipation, headache, increased liver enzymes (AST and ALT), rash, fever and infection.

Inotuzumab Ozogamicin

Inotuzumab Ozogamicin carries the same toxin as gemtuzumab in gemtuzumab ozogamicin, but the MAB component of the ADC is directed against the CD22 antigen on (precursor) B-ALL cells. Safety and treatment emergent adverse events are similar to those of gemtuzumab ozogamicin.

Brentuximab Vedotin

Brentuximab Vedotin is directed against the CD30 antigen, expressed on classical Hodgkin’s Lymphoma Reed-Sternberg and anaplastic large cell lymphoma cells and, in part, mycosis fungoides. It is expressed in embryonal carcinomas, but not in seminomas and is thus a useful marker in distinguishing between these germ cell tumors. The toxin in this ADC is monomethyl auristatin E (MMAE), a dolastatin 10 derivate, derived from peptides occurring in marine shell-less mollusk Dolabella auricularia (Blunt-end Sea Hare) (Fig. 23.11). MMAE is a synthetic drug with a comparable mechanism of action as the taxanes. It inhibits cell division by blocking the polymerization of tubulin. MMAE is 100–1000 times more potent than doxorubicin.

Figure 23.11

MMAE and the attached MAB-linker-conjugate. Via a spacer (para-aminobenzylcarbamate), MMAE is attached to a cathepsin cleavable amino acid linker, followed by an attachment group, consisting of maleimide and caproic acid

Brentuximab Vedotin Safety

The drug has a myelotoxic potential with possible grade 3/4 neutropenia, thrombocytopenia and anemia. Neutropenia can result in severe infections and/or opportunistic infections. In the presence of severe renal or hepatic impairment, a higher frequency of ≥grade 3 toxicities and deaths was observed. Besides severe but rare dermatologic reactions including Stevens-Johnson-Syndrome and Toxic Epidermal Necrolysis, a cumulative, peripheral sensory neurotoxicity, has been observed. This may require a delay, change in dose, or discontinuation of treatment.

Brentuximab and ado Trastuzumab (see below) each carry a toxin that, in case of extravasation, could cause skin necrosis like the vinca alkaloids. As mentioned, the antibodies are conjugated with a cleavable or an uncleavable linker to the toxin. Whether cleavable linkers will be lysed in the extravasation area is unclear at the current time. ADCs with an uncleavable liker will be degraded like proteins and after that, the toxin is released. If and how this may happen to a clinically relevant degree in an extravasation area is also unclear at the current time. Limited clinical experience so far suggests that extravasated ADCs may only cause slight skin reactions.

Anti Growth Factor Receptor Antibodies: Anti-EGFR

■ Anti-EGFR-Strategies

The cell membranous receptors of the epidermal growth factor family consist of four related, functionally different members with tyrosine kinase activity, the homologues EGFR1 to EGFR4 (also called HER1 to HER4 or ErbB1 to ErbB4). The inactive monomers homo- (HER1 with HER1) or hetero-dimerize (HER1 with HER2, or -3, or -4) after external ligand binding. Conformational change leads to auto phosphorylation and subsequent signal transduction for diverse cellular functions including proliferation, cell survival—comprising inhibition of apoptosis-, adhesion, and DNA damage repair. In tumors, this pro survival signal transduction is activated permanently, without binding of a physiological, external ligand such as transforming growth factor-alpha (TGFA), heparin-binding EGF-like growth factor (HBEGF), betacellulin (BTC), amphiregulin (AREG), epiregulin (EREG), epigen (EPGN) and neuregulins (Rubin and Yarden 2001; Schneider and Wolf 2009; Singh et al. 2016). Diseases with malignancy associated overexpression of EGFR1 (HER1) are, for example, colorectal carcinomas (CRC), NSCLC, pancreas carcinomas and squamous cell cancers of the head and neck. Her2 is overexpressed by certain breast and gastric cancers. Over-expression is also known to occur in ovarian, (English et al. 2013; Teplinsky and Muggia 2014; Tuefferd et al. 2007) stomach, and aggressive forms of uterine cancer, such as uterine serous endometrial carcinoma (Buza et al. 2014; Santin et al. 2008). HER2 is over-expressed in 30% of salivary duct carcinomas (Chiosea et al. 2015), however, without any therapeutic consequences up to now. Overexpression has to be demonstrated by immunohistochemistry (IHC) or fluorescent in-situ hybridization (FISH) as a prerequisite for the use of anti-EGFR MABs.

Cetuximab

Cetuximab is a chimeric IgG1 antibody composed of the Fv region of a murine anti-EGFR antibody with human IgG heavy and kappa light chain constant regions. It binds to the extracellular domain of EGFR1 with an affinity five to tenfold greater than endogenous ligands, resulting in inhibition of EGFR signaling. Moreover, it also exerts the cytotoxic immune effector mechanisms described in Chap. 8. It is indicated in the treatment of metastatic colorectal cancer as mono- and combination therapy. Cetuximab also binds to a number of EGFR antigen negative tissues, therefore a saturating loading dose of 400 mg/m² has to be given as a first dose and again, if the weekly interval is exceeded. The weekly follow-up dose is 250 mg/m². Many commonly used chemotherapy regimens for CRC, including irinotecan monotherapy or in combination with infusional fluorouracil, like FolFOx or FolFIri, are administered every 2 weeks (FolFIri treatment regimen (=Folinic acid, Fluorouracil, Irinotecan): irinotecan 180 mg/m² day 1; calcium folinate 400 mg/m² day 1; fluorouracil (5-FU) 400 mg/m² day 1 undiluted bolus injection over 5 min; fluorouracil (5-FU) 2400 mg/m² day 1–2 as 48 h as prolonged infusion; FolFOx treatment regimen (=Folinic acid, Fluorouracil, Oxaliplatin): oxaliplatin 100 mg/m² day 1; calciumfolinate 400 mg/m² day 1; fluorouracil (5-FU) 400 mg/m² day 1 undiluted bolus injection over 5 min; fluorouracil (5-FU) 2400 mg/m² day 1–2 as 48 h as prolonged infusion).

The ability to synchronize the administration of cetuximab and concomitant chemotherapy is more convenient for the patients. Dosing of 500 mg/m² every 2 weeks exhibited predictable pharmacokinetics, which were similar to those of the approved weekly dosing regimen. No differences between the weekly and 2-weekly regimen on pharmacodynamics were observed. Efficacy and safety were also similar (Tabernero et al. 2008). Cetuximab is also used in the treatment of certain squamous cell cancers of the head and neck in combination with radiation or in combination with a platin-based chemotherapy.

Panitumumab

Panitumumab is the fully human version of an anti-EGFR1 MAB and shares the same target as cetuximab with slightly different binding affinity and specificity. In contrast to cetuximab, no loading dose is necessary—it is continuously dosed with 6 mg/kg q2w.

Both antibodies are only effective in pan RAS mutation negative tumors (wild type). K- and N-RAS proteins are G proteins (GTPases) downstream of EGFR and components of EGFR signaling, propagating EGFR signaling events. Activating mutations result in constitutive activation without necessity of ligand binding to the EGFR (Fig. 23.12). In other words: an anti EGFR antibody therapy for patients with activating RAS mutations has no therapeutic benefit, but exposes the patient to the risk of anti-EGFR MAB related toxicity.

Figure 23.12

Mutated RAS carcinomas are resistant to anti EGFR antibodies due to constitutively activated downstream signaling and thus growth promotion

Pan RAS tissue testing is mandatory according to clinical guidelines (Van Cutsem et al. 2014) before initiating a cetuximab or panitumumab therapy. The benefit of adding cetuximab to RAS wild type was shown by the pooled analysis of the OPUS and CRYSTAL studies (Bokemeyer et al. 2012).

Approximately up to 10% of CRC tumors also carry a BRAF mutation. RAS mutations and BRAF mutations are usually mutually exclusive (De Roock et al. 2010), so double testing is not necessary. A BRAF mutation is a strong negative prognostic biomarker and patients with a BRAF mutant CRC have a very poor prognosis (Van Cutsem et al. 2011). However, patients with a KRAS wild type but a BRAF mutation benefit from a cetuximab containing chemotherapy (Van Cutsem et al. 2011). Side matters—location of the CRC: it has been observed for years that patients with right sided colorectal cancer had worse outcomes than those with left-sided disease (Brule et al. 2015). Meanwhile, right and left sided tumors are regarded as molecular distinct (Tejpar et al. 2016). This translates into therapeutic consequences. Whereas treatment of patients with right sided tumors should be started with an antiangiogenic component (bevacizumab), patients with left sided tumors benefit from an anti-EGFR treatment (Venook et al. 2016) (Fig. 23.13).

Figure 23.13

Median survival dependent on the tumor location and first-line regimen in colorectal cancer (CRC (based on Venook et al. 2016)). KRAS-WT = KRAS wild type, not mutated. HR hazard ratio

Necitumumab

Necitumumab is an anti-EGFR recombinant human monoclonal antibody of the IgG1κisotype, currently used against NSCLC in combination with cisplatin (75–80 mg/m² on day 1) and gemcitabine 1200–1250 mg/m² on days 1 and 8). It was approved with a statistically significant difference in median overall survival (OS) of 1.6 months, compared to cisplatin and gemcitabine without antibody (11.5 vs 9.9 months). There is a fixed dosing for necitumumab of 800 mg q3w.

Anti-EGFR MAB Safety

Common anti EGFR therapy associated adverse events include acneiform rash, diarrhea, hypomagnesemia, hypocalcemia, and infusion reactions. Most common are papulopustular rash of the upper trunk and face skin (60–90%), dry and itchy skin (12–16%), and resulting microbial infections (38–70%), conditioned by open pustules as portal of entry for germs. Less frequently, pruritus, hair modifications, and paronychial inflammation occur (Holcmann and Sibilia 2015). By now, the mechanisms underlying skin disorders induced by EGFR inhibitors are well understood (Holcmann and Sibilia 2015). Alterations in chemokine and cytokine production in keratinocytes may result in attraction of inflammatory cells. Disturbed keratinocyte differentiation impairs proper formation of tight junctions and physiological barrier function. The barrier defect and reduced expression of antimicrobial peptides result in bacterial infections. Prophylactic—rather than reactive—management of skin reactions for all patients receiving EGFR inhibitors is recommended. Appropriate prophylaxis can effectively reduce the severity of skin reactions and also a stigmatization as a cancer patients, discernible by their skin eruptions. Skin care has the potential to directly benefit patients and their quality of life. Several recommendations and guidelines have been published (assorted samples: Gutzmer et al. 2011; Hofheinz et al. 2016; Lacouture et al. 2011; Potthoff et al. 2011). Skin changes during therapy proceed in three phases. Skin care has to be adapted to these phases

• Phase I: acneiform skin changes

• Phase II: desiccation phase

• Phase III: dry, sensitive skin

During phase III, the skin is very sensitive to sun light. An unnecessary sun exposure should be avoided, long sleeved outer clothing be worn, and the use of sun blockers should be considered. Furthermore, micro traumatization should be avoided. That is:

• No mechanical manipulation of alleged spots

• No hot hair drying

• No use of curlers, especially no tight wrapped curlers

• No long and hot showering or bathing

• No vigorous rubbing with towels (hard to comply in case of additional, tantalizing itching)

• No occlusion for instance with rubber gloves

• No too tight footwear

• Shaving, electric or blade shaving, always causes (unavoidable) micro traumatization.

Skin cracking that is hardly healing and possibly painful can be sealed with instant glue (like cyanoacrylate glue). This can keep these lesions from worsening or becoming infected. Superglue is also supposed to have some local anesthetic properties (Lacouture et al. 2011).

Electrolyte disturbances (Ca²⁺, Mg²⁺) were initially “simply reported” in the SmPC/FPI. However, especially Mg²⁺ loss occurs frequently. The EGFR is also found in the distal part of the collecting duct and other parts of the kidneys, but with a rather high expression in the ascending part of Henle’s loop and in the distal convoluted tubule, where active re-absorption of magnesium ions takes place (~70%).The EGFR regulates the protein TRPM6 (=transient receptor potential cation channel, under family M member 6) (Schlingmann et al. 2007). By blocking this pump with anti EGFR antibodies, magnesium losses develop (Izzedine et al. 2010) (Patients with a loss-of-function germline mutation in the TRPM6-gene suffer from severe, congenital hypomagnesemia). The elimination half-life of cetuximab is approximately 3–7 days. Assuming that the half-life is 3 days and 5 half-lives are necessary to quantitatively eliminate a drug to have no residual pharmacodynamic activity, then one can be assume that the EGFR dependent magnesium pump is more or less permanently inhibited during the weekly dosing schedules. The same applies for panitumumab with its two-weekly schedule and a half-life of 7.5 days. As a consequence, cumulative hypomagnesaemia of grade 3/4 can develop. This is termed Magnesium Wasting Syndrome (MWS) . The (randomly) observed frequency of MWS ranges between 27% (Fakih et al. 2006) to 36% (Schrag et al. 2005), although in a prospective cohort study 97% of the patients were affected (Tejpar et al. 2007). Besides the known symptoms of magnesium loss such as excitability and cardiac effects, a severe fatigue syndrome can be observed. To detect a MWS early, magnesium levels have to be monitored from baseline (as comparative value), because a loss can occur up to 8 weeks after completing therapy. A hypomagnesemia grade 1/2 can be treated by oral substitution. A grade 3/4 hypomagnesemia has to be treated by parenteral substitution. Fakih et al. recommend 6–10 g (!) magnesium sulfate daily or twice weekly (Fakih 2008). This magnitude of necessary substitution was confirmed for cetuximab (Schrag et al. 2005) as well as for panitumumab induced MWS (Cheng et al. 2009). Such an amount of magnesium sulfate has to be given as a protracted infusion, because of a quick compensatory elimination from short infusions by the kidneys. Ambulatory pumps such as those for infusional 5-FU can be used. In contrast to anti-EGFR MABs, no magnesium deficiency—not to mention MWS—has been reported with the use of small molecular inhibitors EGFR tyrosine kinase inhibitors such as erlotinib or gefitinib, except in context with diarrhea. Altundag et al. (2005) argue that the presence of magnesium stearate as a tableting ingredients compensates the drug induced deficiency. However, regarding the doses needed for supplementation, this seems rather unlikely. During a phase I study with erlotinib combined with the multikinase inhibitor sorafenib, phosphate deficiencies were observed (Izzedine et al. 2010). MWS seems to be a class effect of anti EGFR MABs. Detected relatively late after approval of cetuximab and panitumumab, a frequency for hypomagnesemia of 83% is stated in the FPI/SmPC of necitumumab. Meanwhile, electrolyte monitoring (potassium, calcium, magnesium) is recommended to prevent cardiopulmonary arrest.

Hypersensitivity risk by tick bite : an unexpected connection with cetuximab’s structure. As with other MABs, hypersensitivity prophylaxis by premedication with an antihistamine is recommended. European product specifications also recommend the use of a corticosteroid premedication. However, severe anaphylaxis during the first exposure to cetuximab has been observed. These reactions to cetuximab developed rapidly and symptoms often peaked during or within 20 min following the first infusion of the antibody and occasionally proved fatal. Many of the affected patients also reported to be intolerant to mammalian (red) meat such as beef and pork. These first events were associated with a specific geographical region: a group of southern US states. The highest occurrence of these hypersensitivity reactions was observed in Virginia, North Carolina, Tennessee, Arkansas, Oklahoma, and Missouri (Chung et al. 2008). Subsequently, it has become clear that the syndrome of delayed anaphylaxis to red meat is also most common in these same states (Commins et al. 2009). The similarity between the region for reactions to cetuximab and the maximum incidence of Rocky Mountain spotted fever suggested that tick bites might be relevant to these reactions.

Subsequently, it could be shown that tick bites can induce an immunological reaction against the oligosaccharide galactose-alpha-1,3-galactose (alpha-gal), most likely through components in the tick saliva. Alpha-gal is naturally not present in humans. Immunocompetent individuals may form antibodies to alpha-gal, that makes alpha-gal an immunogenic carbohydrate. The alpha-gal oligosacharide is also present at a glycosylation site (Asn88) on the Fab region of the heavy chain of cetuximab, a chimeric IgG1 with murine components. Each cetuximab molecule contains two alpha-gal epitopes that can cross-link the high affinity receptor for IgE on mast cells leading to mast cell activation and release of hypersensitivity mediators (Saleh et al. 2012) (Fig. 23.14). IgE binding to alpha-gal was later linked to allergic reactions to red meat in America and Europe.

Figure 23.14

Infusion reactions with cetuximab are linked to the presence of IgE antibodies directed against the alpha-gal component of the Fab fragment of the cetuximab heavy chain

Despite the development of an ELISA test for anti-cetuximab IgE for the identification of patients with high risk (Mariotte et al. 2011), no recommendations for pre-therapeutic testing exist up to now. The phenomenon, however, is mentioned in the SmPC. For further reading consult (Chung et al. 2008; Commins et al. 2009; Saleh et al. 2012; Berg et al. 2014; Commins et al. 2011; Steinke et al. 2015).

■ Anti Growth Factor Receptor Antibodies Anti-EGFR2 (=Anti HER2/NEU)

Trastuzumab

Trastuzumab is mainly indicated for HER2/neu overexpressing breast cancers in different clinical situations, as monotherapy as well as in combination therapy. It is also approved for HER2/neu overexpressing gastric cancers in combination with chemotherapy. Overexpression has to be verified by IHC or FISH testing. Dosing depends on the interval. For the weekly regimen, a 4 mg/kg loading dose is necessary, followed by 2 mg/kg weekly. The loading dose for the 3 weekly interval is 8 mg/kg followed by 6 mg/kg. Like rituximab, a fixed dose preparation of s. c. trastuzumab is available (in Europe). 600 mg are given q3w over 2–5 min in the thigh. No loading dose is necessary. However, this time-saving procedure compared to i.v. infusions is counteracted by the EMA’s demand to supervise the patients for 6 h (!) after the first and for 2 h after subsequent injections.

Pertuzumab

Pertuzumab is another anti HER2 antibody. Trastuzumab binds to the HER2 subdomain IV and disrupts ligand-independent HER2 signaling (Fig. 23.15). Pertuzumab binds to subdomain II and blocks ligand-dependent HER2 heterodimerization with HER1, HER3, and HER4. Therefore, pertuzumab has been classified as the first HDI = HER2 dimerization inhibitor . The combination of pertuzumab and trastuzumab significantly augmented anti-tumor activity in HER2-overexpressing xenograft models, which was the rationale for the development of this double antibody based anti-Her2 strategy. Improved anticancer activity was proven in patients treated with pertuzumab in combination with trastuzumab compared to either drug alone (Baselga et al. 2012; Cortes et al. 2012).

Figure 23.15

(a) HER2/3 heterodimerization with subsequently the strongest mitogenic signaling and activation of two key pathways that regulate cell survival and growth (b) Pertuzumab binds to subdomain II and blocks ligand-dependent HER2 heterodimerization with HER1, HER3, and HER4. (c) Trastuzumab binds to subdomain IV and disrupts ligand-independent HER2 signaling. Trastuzumab in combination with pertuzumab provide a more comprehensive blockade of HER2-driven signaling pathways

Pertuzumab is given as a combination therapy with trastuzumab on a 3 weekly basis (with docetaxel). The initial loading dose is 840 mg followed by 240 mg q3w (Trastuzumab: 8 mg/kg loading dose followed by 6 mg/kg q3W).

Ado-Trastuzumab-/Trastuzumab Emtanside

Ado-Trastuzumab-/Trastuzumab Emtanside (TDM-1 ) is the third anti-HER2 antibody, an ADC coupled with a toxin. This mertansinoide is a 19 unit lactam of the ansamycine type, derived from an Ethiopian bush of the Maytenus genus (Maytenus serrata) (Fig. 23.16). The toxin acts like the vinca alkaloids as a mitosis inhibitor. Dosing is based on body weight with 3.6 mg/kg q3w.

Figure 23.16

The antibody-toxin-construct in TDM-1

Anti-HER2 MAB Safety

The Her2 receptor has essential roles in embryonal and fetal development and tissue protection, i.e. via anti apoptosis. Particularly neuronal and non-neuronal tissues, including cardiac myocytes, require Her2 for normal development (Negro et al. 2004; Zhao et al. 1998). By blocking these protective functions, the corresponding toxicity, namely cardiotoxicity, is observed. This applies to all three anti-HER2 MABs. All of them carry a warning message in their FPI/SmPC for cardiomyopathy and/or the development of a reduced left ventricular ejection function. That means all anti-HER2 antibodies have the potential to cause ventricular dysfunction and congestive heart failure. There is a higher risk for patients who receive anthracyclines, taxanes or cyclophosphamide in combination and/or during previous therapy. An evaluation of left ventricular function in all patients prior to and during treatment with these MABs is mandatory. Post approval surveys revealed a potential for lung toxicity, including interstitial lung disease. Some (rare) cases developed respiratory distress syndrome, including some with fatal outcome. Exacerbation of chemotherapy-induced neutropenia can occur with trastuzumab, and can also occur when combining pertuzumab and trastuzumab. For TDM-1, hepatotoxicity, has been reported, predominantly as asymptomatic elevations of transaminases, and neurotoxicity. This is most probably attributable to MMAE. The most common adverse reaction are infusion reactions (usually mild to moderate), which rarely require discontinuation of therapy. A routine prophylaxis with antihistamines, antipyretics and/or corticosteroids is not recommended. Skin toxicity like rash, pruritus and dry skin is described. However an intense prophylaxis therapy as required for the anti-EGFR MABs- is apparently not necessary.

Anti-Angiogenic Antibodies

A small (undetectable) tumor is nourished by passive diffusion. When it has reached dimensions of 1 to 2 mm across, diffusion is not sufficient any longer. This leads to evolutionary pressure for the tumor and, in the sense of gain-of-function, it can secrete autocrine pro angiogenic growth factors such as VEGF (Vascular Endothelial Growth Factor). The angiogenic switch has been turned on. The subsequent formation of new blood vessels from pre-existing vessels (angiogenesis) is crucial for tumor development, growth and metastasis (Ohta et al. 1996). VEGF, with its different subtypes (VEGF-A –E), is a secretory, proangiogenic cytokine and one of the major regulators of the process of neovascularization (Ohta et al. 1996). An important regulating process in angiogenesis is the interaction of the VEGF subtypes with their receptors. Fig. 23.17 shows the members of the VEGF family with their receptors and the resulting effects.

Figure 23.17

VEGF-family and their corresponding receptors. PlGF was discovered in human placenta and shares a 50% homology to VEGF-A. VEGF vascular endothelial growth factor, PlGF placenta growth factor, VEGF-R vascular endothelial growth factor receptor, NRP neuropilin

Anti-angiogenic therapy leads to:

• Regression of existing microvasculature, known as antivascularisation

• Normalization of the surviving vasculature offering optimal chemotherapy delivery within the tumor, thus enhancing antitumor properties

• Antiangiogenic effect leading to the inhibition of new vessel growth, offering improved response rates and tumor death with the potential of improving patient relevant outcomes (progression free survival—PFS, time to progression—TTP, overall survival—OS).

Up to now, there are two principles of anti-angiogenic therapy.

• 1. Intercepting the soluble growth factors in the peripheral blood before they can interact with their receptors (bevacizumab, aflibercept)

• 2. Blocking the receptor a) by a MAB (ramucirumab) from the extracellular space, b) by a small molecule kinase inhibitor from the intracellular space (examples: axitinib, cabozantinib, nintedanib, pazopanib, regorafenib, sorafenib, sunitinib—not further discussed here).

■ Bevacizumab

Bevacizumab is directed against VEGF-A and its isoforms. VEGF is intercepted in the peripheral blood and tumor microenvironment, and thus neutralized before it can exert proangiogenic effects. Bevacizumab is used against several solid tumors like CRC (initial approval), NSCLC, breast- and renal cancer. Dosing depends on the schedule interval and the underlying disease. 5–10 mg/kg q2w or 7.5–15 mg/kg q3w are given.

■ Ziv-aflibercept

Ziv-aflibercept / Aflibercept (in EU) is a recombinant fusion protein consisting of the VEGF binding extracellular domain of human VEGF-receptors 1 and 2, fused to the Fc-part of human IgG1. It binds to all VEGF-A isoforms with a 100-fold higher affinity than bevacizumab. It also binds to VEGF-C and -D and to PlGF. It is designated as VEGF trap. The clinical use in oncology is restricted to CRC in combination with FolFIri at a dose of 4 mg/kg q2w. A non-oncologic indication of aflibercept (of note the INN) is the treatment of patients with neovascular (wet) age-related macular degeneration by ophthalmic intravitreal injection.

■ Ramucirumab

Ramucirumab is a recombinant human IgG1 monoclonal antibody that specifically binds to vascular endothelial growth factor receptor 2. Its clinical use comprises NSCLC, gastric, and colorectal cancer as single agent (gastric or gastro-esophageal junction adenocarcinomas) or in combination with selected chemotherapy regimens. The dosage for gastric cancer/CRC is 8 mg/kg qw2, whereas for NSCLC it is 10 mg/kg q3w.

Anti-Angiogenic MAB Safety

Initially it was assumed that VEGF-targeted therapies would be toxicity free. However, clinical trials and experience revealed a number of adverse events associated with anti-angiogenic agents, MABS as well as small kinase inhibitors, which can be summarized as a class effect (Chen and Cleck 2009; Hutson et al. 2008). These treatment-emergent adverse events comprise:

• Hypertension

• Impaired wound healing

• Haemorrhage, including severe courses

• Proteinuria

• Nephrotic syndrome or thrombotic microangiopathy

• Gastrointestinal perforation

• Fistula formation

• Arterial thromboembolic events

• Grade 4 venous thromboembolic events (including pulmonary embolism)

• Posterior reversible encephalopathy syndrome (PRES), also known as reversible posterior leukoencephalopathy syndrome (RPLS), colloquially referred to as “cortical blindness”.

Pre-therapeutic hypertension has to be adjusted and monitored through therapy. Sometimes, “aggressive” interventions for blood pressure control have to be undertaken or therapy has to be discontinued. Blood pressure is mainly driven by the VEGF-R2, e.g. by NO and prostacyclin I₂ release from endothelial cells. By blocking VEGF-R2 and intercepting its ligands, the synthesis of vasodilators is suppressed, which in turn increases the peripheral resistance and in consequence the blood pressure (Verheul and Pinedo 2007). Elevated peripheral vascular resistance can also contribute to a reduced left ventricular ejection function and thus be the cause for (congestive) heart insufficiency.

Wound healing depends on neoangiogenesis. Therefore, it is easy to understand that wound healing is impaired by anti-angiogenic therapy. The mentioned MABs may be used at the earliest 4 weeks after surgery and after complete wound healing. They also have to be interrupted for 4 weeks before a planned surgery. Gastrointestinal perforations, especially along surgical sutures, have occurred.

PRES or RPLS is a rare (<1%) but severe side effect of anti-angiogenic or blood pressure elevating drugs (examples of the latter: proteasome inhibitors). RPLS is a brain-capillary leak syndrome related to hypertension, fluid retention, and the cytotoxic effects of immunosuppressive agents on the vascular endothelium. It seems that severe hypertensive encephalopathy leads to RPLS and vasogenic edema of the posterior cerebral white matter, induced by endothelial dysfunction and a disrupted blood–brain barrier (Verheul and Pinedo 2007; Widakowich et al. 2007). This rare syndrome regained attention after the approval of bevacizumab, but for most of the anti-angiogenic kinase inhibitors this side effect has been described, as well as for proteasome inhibitors such as carfilzomib, which also elevates blood pressure. Patients who experience a RPLS must never be exposed again to the causative drug.

Immune Oncology

■ Immune Agonistic Antibodies

In principle, the human immune system can prevent tumorigenesis. Tumor cells, however, have the capability to escape the immune system (Vesely et al. 2011). (Re-)activating the immune system to eliminate cancer cells to produce clinically relevant responses has been a long-standing “dream of mankind”. T cells have an important role to play in fighting cancers. To avoid collateral damage of host tissues and organs, T cell action is balanced by inhibitory signals and molecules—the so-called immune checkpoints. They are critical for maintaining self-tolerance on the one hand and modulating the duration and amplitude of immune responses on the other.

Normally, after T-cell activation, CTLA4 (Cytotoxic T-Lymphocyte Antigen 4; CD152), is upregulated on the plasma membrane. Its functions is to downregulate T-cell activity through a variety of mechanisms, including preventing co-stimulation by outcompeting CD28 for its ligand, B7, and also by inducing T-cell cycle arrest. Through these mechanisms and others, CTLA-4 has an essential role in maintaining normal immunologic homeostasis. CTLA4 downmodulates the amplitude of T cell activation. Blockade of CTLA4 by MABs such as ipilimumab keeps T cells stimulated (Fig. 23.18).

Figure 23.18

Simplified mechanism of action of anti CTLA4 antibodies. Left: T cell activation by antigen presenting to the T cell receptor with the co-stimulatory signal B7 to CD28. Middle: Under physiological conditions, the activated T cells are downregulated after 48–72 h by the displacement of costimulatory CD28 with CTLA4. The B7-CTLA4-axis slows down the T cells up to complete inhibition and maintains self-tolerance in the periphery. Right: By blocking CTLA4 with MABs, the T cell remains activated, attacking tumor tissues. (APC antigen presenting cell, CTLA4 cytotoxic T-lymphocyte antigen 4, MHC major histocompatibility complex, TCR T cell receptor)

In contrast to CTLA4, which primarily regulates the amplitude of the early stages of T cell activation, the major role of PD1 is to limit the T cell activity in peripheral tissues. Despite its name—programmed cell death protein-1—PD1 does not induce cell death directly. When engaged with one of its ligands, PD-L1 or PD-L2, PD1 inhibits kinases that are involved in T cell activation. In summary, this pathway is a “stop signal” for T cells. Tumors use these inhibitory pathways to evade an immune attack through overexpression of PD-L1. As a result, this blocks the generation of an immune response to the tumor (cf. Figs. 23.19 and 23.20).

Figure 23.19

Naive T cells are activated by the presentation of tumor antigens. During this priming phase, PD-1 is upregulated as a physiological reaction to protect unaffected host tissue. The corresponding ligand, PD-L1, expressed on tumor cells, downregulates T cells, pretending to be “friendly”, healthy tissue. The magnifier shows the details of the T cell receptor (TCR) interaction with tumor antigen presenting Major Histocompatibility Complex (MHC). Inactivating programmed cell death receptor 1 (PD-1) is stimulated by the corresponding ligands (PD-L1/2), resulting in T cell exhaustion. (MHC major histocompatibility complex, PD-1 programmed cell death receptor1, PD-L1/2 PD1/2-ligand, TCR T cell receptor)

Figure 23.20

Anti PD-1 and/or anti PD-L antibodies prevent the inactivation of immune competent T cells

Long-term exposure to antigens in the presence of inflammatory cytokines induces a distinct phenotype in T cells, characterized by loss of effector functions, sustained expression of inhibitory receptors, poor proliferative capacity and decreased cytotoxic functions. This progressive loss of T-cell effector functions is commonly seen during chronic viral infections and in cancer. It is termed “T-cell exhaustion”. T cells detect the tumor, accumulate in the tumor microenvironment, however, are silenced by inhibitory molecules expressed on the tumor surface such as PD-L1/2. PD-1/PD-L1 inhibitors pharmacologically prevent the PD-1/PD-L1 interaction, thus facilitating a positive immune response to kill the tumor. For further immune oncology (IO) information, the reader is referred to Alsaab et al. (2017), Ferris (2013), Intlekofer and Thompson (2013), Li et al. (2016), Pardoll (2012), Rotte et al. (2018), Suzuki et al. (2016)).

Immune oncology MABs do not show a clear dose-response relationship. They are either dosed by body weight or with a fixed dose. For the future it may be possible that antibodies currently dosed by body weight may be switched to fixed dosing.

Due to their mechanism of action, infiltrating the tumor tissue followed by tumor cell killing, an initial increase of tumor lesions was observed. From the formal point of view, these patients were RECIST progressive (RECIST = Response Evaluation Criteria In Solid Tumors). However, it would have been a mistake to stop treatment too early, as biopsies confirmed inflammatory cell infiltrates with an apparent enlargement of the tumor. This phenomenon is termed pseudoprogression or “tumor flare” (short overview Chiou and Burotto 2015). Thus, to ascertain a clinical benefit, it can take 12 weeks or more after the first infusion and may include the emergence of (temporary) new lesions. To avoid misclassification according WHO in immune oncology, immune-related response criteria (irRC) were developed in 2009 (Wolchok et al. 2009).

■ Anti CTLA4 Antibodies

Ipilimumab

Ipilimumab is the first anti CTLA4 , recombinant human monoclonal antibody. It is used as monotherapy for the treatment of melanoma in different clinical situations. 3 mg/kg q3w for 4 doses are used for unresectable or metastatic melanoma. In a certain adjuvant setting (cutaneous melanoma with pathologic involvement of regional lymph nodes of more than 1 mm that have undergone complete resection, including total lymphadenectomy), 10 mg/kg q3w for 4 doses followed by 10 mg/kg q12w up to 3 years are recommended. Fig. 23.18 illustrates the simplified mechanism of action.

■ Anti PD1- and Anti PD-L1 Antibodies

While development of new anti PD1/anti PD-L1 antibodies is rapidly progressing and will likely provide new agents, the following MABs were on the market at the time of the creation of this manuscript (Table 23.2):

Table 23.2 Approved anti PD-1 and anti PD-L1 antibodies, effective April 2018

The anti PD1/anti PD-L1 antibodies are used against different solid tumors. Depending on the underlying disease and the clinical situation (first-line treatment or relapsed tumor), PD-L1 testing is sometimes mandatory. The relevance and usefulness of PD-L1 as a predicting biomarker is still under debate. Fig. 23.19 illustrates the pathological mechanisms, Fig. 23.20 the pharmacodynamic intervention with MABs.

Immune Oncology MAB Safety: Playing with Fire?

Due to their mechanism of action , immune checkpoint inhibitors in the form of MABs against CTLA-4, PD-1, and PD-L1, have a unique spectrum of toxicity that differs from the typical adverse events seen with chemotherapeutic agents. By inhibiting checkpoint molecules, the immune system is activated or even over-activated or, from the viewpoint of the normal tissue, deregulated (non-recognizing “self”). These new kind of autoimmune-like symptoms are based on the loss of self-tolerance and termed immune-related adverse events (irAEs) . For CTLA-4–blocking antibodies, toxicities seem to be dose related, because the rate of grade 3 to 4 drug-related serious irAEs increased from 5% to 18% when the dose was increased from 3 to 10 mg/kg whereas it was 0% at a dose of 0.3 mg/kg. In contrast, the toxicities related to PD-1 blockade are similar at doses ranging from 0.3 to 10 mg/kg, exemplary observed with nivolumab. The observed toxicities depend on

• The patient population/the underlying disease

• The dose (especially for CTLA-4 antibodies)

• The schedule

That means, the same dose or schedule in different diseases might result in different toxicity profiles. Combinational checkpoint blockade can result in an increase of grade 3/4 treatment emergent adverse events (TEAE) up to 53% (Wolchok et al. 2013). The toxicity pattern is based on induced autoimmunity and comprises:

• Skin irAEs, including Stevens-Johnson-Syndrome and Toxic Epidermal Necrolysis (both rare)

• Gastrointestinal irAEs—autoimmune colitis

• Autoimmune hepatitis

• Autoimmune pancreatitis

• Autoimmune thyroiditis

• Autoimmune hypophysitis

• Neurological irAEs

• Autoimmune pneumonitis

• Ocular irAE (rare), like autoimmune uveitis and autoimmune episcleritis

In December 2016 the Federal Institute for Drugs and Medical Devices, Germany, informed the medical professional circles about suspected cases of pancytopenia/agranulocytosis. Onset was between 12 and 274 days after start of therapy. Three instances were fatal. Another rare irAE was myocarditis (9 cases), with two of them having an additional myositis and rhabdomyolysis. Four cases had a fatal outcome. In principle, every tissue and organ can be affected (Fig. 23.21).

Figure 23.21

Affected tissues organs and symptoms by autoimmune related adverse reactions. Listing is not intended to be exhaustive

Toxicity/iRAE Management

Physicians and pharmacists should know about and be able to recognize irAE as such and the recommended interventions (Davies and Duffield 2017; Haanen et al. 2015; Weber et al. 2012; Weber et al. 2015).

In severe diarrhea with associated signs of colitis, the usual counter-measures (fluid + electrolytes ⇨ loperamide ⇨ budesonide) are insufficient. Intravenous or oral steroid therapy has to be initiated. For patients in whom intravenous steroids followed by high-dose oral steroid therapy does not lead to initial resolution of symptoms within 48–72 h, treatment with infliximab at 5 mg/kg can be used as an “emergency brake”. Once relief of symptoms is achieved with infliximab, it should be discontinued. A prolonged steroid taper over 45–60 days should be instituted. There may be a waxing and waning of the GI adverse effects. As steroids are tapered, there can be a recrudescence of symptoms, mandating a retapering of steroids starting at a higher dose, a more prolonged taper, and the (re-) use of infliximab. Prophylactic use of budesonide cannot be recommended, based on a phase II study (Wolchok et al. 2010).

Autoimmune hepatitis is likewise treated with (high dose) corticosteroids. If serum transaminase levels do not decrease within 48 h after initiation of systemic steroids, oral mycophenolate mofetil 500 mg q12h should be considered. Infliximab has to be avoided, because of its potential own hepatotoxicity. As described above, resurgence of symptoms and steroid tapering is necessary.

Difficult in diagnosis can be the auto hypophysitis with symptoms of headache, nausea, vertigo, behavior change, visual disturbances such as diplopia, and weakness. In this context new occurrence of brain metastases have to be excluded. Baseline measurement of pituitary, thyroid, adrenal, and gonadal status, i.e. serum morning cortisol, adrenocorticotropic hormone [ACTH], free triiodothyronine [T3], free thyroxine [T4], thyroid-stimulating hormone [TSH], testosterone in males and follicle-stimulating hormone, luteinizing hormone, and prolactin in females is recommended.

Immune-related pancreatitis generally manifests as an asymptomatic increase of amylase and lipase. Some patients experience more or less unspecific symptoms like fevers, malaise, nausea and vomiting and/or abdominal pain. As described for other irAEs, a steroid taper is indicated, but often this has minimal immediate effects. The symptoms of an autoimmune pancreatitis resolve slowly.

The irAEs caused by checkpoint blockade exhibit a characteristic pattern in the timing of their occurrence, shown in Fig. 23.22 (Weber et al. 2012).

Figure 23.22

irAEs exhibit a characteristic pattern in the timing of their occurrence. The frequency is not considered (from Weber et al. 2012)

After 2–3 weeks of initiation of a checkpoint blocker therapy, skin-related irAEs can be expected. Gastrointestinal side effects emerge after approximately 5–6 weeks. Hepatic irAEs occur after 6–7 weeks, and endocrinological irAEs (autoimmune hypothyroiditis or hypophysitis) after an average of 9 weeks. Long-term effects of immune oncology therapy are currently still largely unknown.

Self Assessment Questions

■ Questions

• 1. Against what kind of molecular targets can MABs be directed? Give examples

• 2. By which factors can the pharmacokinetics of MABs be influenced?

• 3. What stands ADC for?

• 4. Why is Rituximab in CLL dosed with 375 mg/m² initially but escalated to 500 mg/m²in subsequent cycles?

• 5. Describe the differences in clinical effects between type I and type II anti-lymphoma antibodies.

• 6. How many subtypes of the epidermal growth factor receptors exist and how are they designated?

• 7. Which diagnostic actions are mandatory before using anti-EGFR/anti-Her2 antibodies?

• 8. Does it make therapeutically a difference if a colorectal carcinoma is right sided or left sided?

• 9. What is the main difference in the mechanism of action between ‘classical’ antibody treatment of cancers and immune agonistic antibodies?

• 10. Describe the toxicity profile of checkpoint inhibitors.

■ Answers

• 1. MABs can be directed against

  • Surface antigens like CD antigens (CD20, CD30)

  • Receptors (EGFR)

  • Growth factors (VEGF)

  • Activating immune checkpoints on T cells (PD1)

  • Suppressing immune checkpoints on tumor cells (PD-L1)

• 2. Tumor burden which corresponds to the amount of target antigen.

  • Neutralizing anti-MAB-antibodies

  • Gender

  • Age

• 3. ADC is the abbreviation for antibody drug conjugate, where an antibody is chemically linked to a toxin with anti-tumor activity. The toxins are too toxic to be given in unconjugated form.

• 4. In CLL the CD20 antigen density is lower than in other CD20 positive lymphomas. The reduced dose for the first infusion is necessary because of a usually high tumor load in need for therapy in CLL blast crisis. Responding to rituximab therapy bears a high risk of tumor lysis and cytokine release syndrome as well as other infusion related reactions.

• 5. Type I antibodies cause predominant CDC, type II antibodies ADCC

• 6. 4 subtypes, EGFR 1–4 or ErbB 1–4 or Her 1–4

• 7. Verification of EGFR or Her2 overexpression, in case of Her2 with IHC or FISH. Pan-RAS wild type testing for cetuximab and panitumumab.

• 8. Depending on the localization, these tumors are regarded as biologically different. Patients with a right sided tumor benefit from an antiangiogenic drug containing first line therapy, whereas left sided tumors benefit from an anti EGFR strategy if they are not RAS mutated.

• 9. The so-called check point inhibiting antibodies do not attack the tumor itself, they restore their immunogenicity and/or (re-)activate immune competent cells.

• 10. The toxicity pattern consists of loss of self-olerance, resulting in autoimmune reactions (immune related adverse events) such as

  • Skin irAEs, including Stevens-Johnson-Syndrom and Toxic Epidermal Necrolysis (both rare)

  • Gastrointestinal irAEs—auto immune colitis

  • Autoimmune hepatitis

  • Autoimmune pancreatitis

  • Autoimmune thyreodits

  • Autoimmune hypophysitis

  • Neurological irAEs

  • Autoimmune pneumonitis

  • Ocular irAE (rare), such as autoimmune uveitis and autoimmune episcleritis