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Recent Results in Cancer Research
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Matthias Theobald Editor

Current
Immunotherapeutic
Strategies in Cancer
Recent Results in Cancer Research

Volume 214

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Matthias Theobald
Editor

Current
Immunotherapeutic
Strategies in Cancer

123
Editor
Matthias Theobald
Department of Hematology, Oncology
and Pneumology, University Cancer
Center (UCT) Mainz
Johannes Gutenberg University
Medical Center
Mainz, Germany

ISSN 0080-0015 ISSN 2197-6767 (electronic)

Recent Results in Cancer Research
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Contents

Current Development of Monoclonal Antibodies

in Cancer Therapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
Sagun Parakh, Dylan King, Hui K. Gan and Andrew M. Scott
Clinical Experience with Bispecific T Cell Engagers . . . . . . . . . . . . . . . 71
Nicola Gökbuget
Advances and Challenges of CAR T Cells in Clinical Trials . . . . . . . . . 93
Astrid Holzinger and Hinrich Abken
Targeting Cancer with Genetically Engineered TCR T Cells . . . . . . . . . 129
Thomas W. Smith Jr. and Michael I. Nishimura
Personalized Neo-Epitope Vaccines for Cancer Treatment . . . . . . . . . . . 153
Mathias Vormehr, Mustafa Diken, Özlem Türeci, Ugur Sahin
and Sebastian Kreiter
The Era of Checkpoint Inhibition: Lessons Learned
from Melanoma . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 169
Annette Paschen and Dirk Schadendorf

v
Current Development of Monoclonal
Antibodies in Cancer Therapy

Sagun Parakh, Dylan King, Hui K. Gan and Andrew M. Scott

1 Antibody Structure

Antibodies are the epitome of specificity with an estimated ten billion different
antibodies produced by human B cells; there is an extraordinarily diverse range of
antibodies capable of being produced by the immune system (Fanning et al. 1996).
Antibodies are made up of four polypeptide chains, two identical light chains and
two identical heavy chains, which are joined by disulphide bridges forming a
structure that is similar to the shape of a Y (Fig. 1) (Merino 2011). Both the light
and heavy chains are comprised of variable and constant domains, each with dif-
fering functions (Merino 2011). The variable domains determine antigen specificity,
and the constant domains determine immunoglobulin (Ig) class. For the light
chains, the constant domain differs depending on whether they are encoded by j or
k genes (Merino 2011). Similarly, the constant domain of the heavy chain varies
with 5 genes (c, µ, a, d and e), and this determines the overall antibody class (IgG,
IgM, IgA, IgD and IgE, respectively) (Merino 2011). Furthermore, IgA has two

S. Parakh
D. King
H. K. Gan
A. M. Scott (&)

Tumour Targeting Laboratory, Olivia Newton-John Cancer Research Institute,
145 Studley Road, Heidelberg, Melbourne, VIC 3084, Australia
e-mail: [email protected]
S. Parakh
H. K. Gan
Department of Medical Oncology, Olivia Newton-John Cancer and Wellness Centre,
Austin Health, Heidelberg, Melbourne, Australia

S. Parakh
D. King
H. K. Gan
A. M. Scott

School of Cancer Medicine, La Trobe University, Melbourne, Australia
Department of Molecular Imaging and Therapy, Austin Health, Melbourne, Australia
Department of Medicine, University of Melbourne, Melbourne, Australia

© Springer Nature Switzerland AG 2020 1

M. Theobald (ed.), Current Immunotherapeutic Strategies in Cancer,
Recent Results in Cancer Research 214,
https://doi.org/10.1007/978-3-030-23765-3_1
2 S. Parakh et al.

Fig. 1 Antibody structure: Antibodies are made up of four polypeptide chains, two identical
light chains and two identical heavy chains, joined by disulphide bridges. Heavy chains comprise
one variable (VH) domain followed by a constant domain (CH1), a hinge region and two more
constant (CH2 and CH3) domains. The light chain has one variable (VL) and one constant (CL)
domain. The two arms in the Y-shaped structure contain the antigen-binding sites, the fragment
antigen-binding (Fab) region, along with the base of the Y-shaped structure called the fragment
crystallizable (Fc) region. Antigen specificity in the Fab region is determined by
complementarity-determining regions (CDRs) within the variable domains

subclasses, IgA1 and IgA2, and IgG, four: IgG1, IgG2, IgG3 and IgG4 (Merino
2011). In healthy people, IgG antibodies represent approximately 75% of serum
antibodies, 15% are IgA, 10% are IgM, along with very small amounts of circu-
lating IgD and IgE antibodies. IgG antibodies are the primary isotype used in cancer
therapy and as such will be the major focus in the following sections.
Functionally, antibodies are divided into two parts; the two arms in the Y-shaped
structure contain the antigen-binding sites and are named as the fragment
antigen-binding (Fab) region, along with the base of the Y-shaped structure which
mediates immunological signalling by antibodies and is called the fragment crys-
tallizable (Fc) region. The Fab arm of an IgG antibody contains the full light chains
and part of the heavy chain, each with their own constant and variable domains.
Antigen specificity in the Fab region is determined by complementarity-
determining regions (CDRs) within the variable domains. These CDRs have the
greatest sequence variation within antibodies, and this feature gives rise to the
Current Development of Monoclonal Antibodies in Cancer Therapy 3

diverse range of antigen specificities. There are three CDRs for each variable
region, which means six CDRs (heavy and light) for each Fab arm and twelve in
total for a single antibody molecule. The six CDRs on each Fab arm fold together to
form the antigen-binding pocket, and this allows an antibody to be able to simul-
taneously bind two epitopes. When antibodies recognize a soluble antigen, this
simultaneous binding can produce large multimeric structures called immune
complexes.
Within the immune system, a principle function of antibodies is to neutralize
pathogens such as bacteria and viruses. The CDRs within the variable regions of an
antibody recognize a specific molecular structure of an antigen, called the epitope,
present on the pathogen. Because of the random nature of antibody generation in
the development of each individual B cell, there are millions of B cells circulating at
any given time that each recognize a different antigen. Once a B cell encounters an
invading pathogen with its unique epitope, it undergoes maturation with the help of
specific T cells and produces large amounts of soluble antibody. Multiple B cells
will recognize different epitopes present on the pathogen, and so many different
antibodies will be produced. Once produced, these antibodies bind their antigen on
the surface of the bacteria or virus to neutralize the pathogen and mark it for
destruction by innate immune effector cells.

1.1 Target Antigens

Following the discovery of antibodies and their functions, it was realized that they
would be potentially efficacious for the treatment and diagnosis of cancers (Rettig
and Old 1989; Scott et al. 2012). Because antibodies are uniquely specific for their
target antigen, they could be used to directly target tumours expressing the antigen.
For ideal targeting of tumour-associated antigens (TAA), what is required is a cell
surface antigen on the tumour that is mutated, overexpressed or selectively
expressed when compared to normal tissue (Scott et al. 2012). Ideally, the target
antigen would be homogenously expressed within the tumour and antigen secretion
would be minimal, in order to reduce antibody trapping in the circulation (Scott
et al. 2012). In addition to expression, antigen function and effect on downstream
signalling are also taken into consideration when selecting a target.
TAAs that are targeted by therapeutic antibodies can be initially grouped on
what type of cancer they target (Tables 1 and 2). Haematological cancers are
usually targeted through cluster of differentiation (CD) antigens that include CD20,
CD30, CD33 and CD52 (Scott et al. 2012), whereas solid tumours can be targeted
through a variety of antigens that fall into different categories based on their
function. The epidermal growth factor receptor (EGFR) is one such example of a
TAA that has been successfully targeted in cancer therapy (Scott et al. 2012).
Antibodies that target EGFR abrogate the native function of the receptor, thereby
inhibiting tumour growth, and can also recruit innate immune cells through
Fc-signalling to mediate killing of the tumour.
 4
Table 1 Approved monoclonal antibodies in solid tumours
Target Drug Indication Tx line Year Trial Treatment arms Endpoints
HER2 Trastuzumab Metastatic breast 1st 1997 Slamon et al. (2001) Chemotherapy + ORR PFS OS
cancer trastuzumab versus 50% versus 7.4 versus 4.6 25.1 versus
chemotherapy 32%; p < months; p < 20.3 months p
0.001 0.001 = 0.046
Node-positive Adjuvant 2006 NSABP B31 + TAC ± trastuzumab DFS OS
breast cancer N9831 (Perez et al. 62% versus 75% versus
2011, 2014) 74%; p < 84%; p < 0.001
0.001
Metastatic gastric 1st 2010 TOGA (Bang et al. Chemotherapy ± ORR PFS OS
or GEJ 2010) trastuzumab 47% versus 6.7 versus 5.5 13.8 versus
adenocarcinoma 35% months; p = 11.1 months; p
0.002 = 0.046
Pertuzumab Metastatic breast 1st 2012 CLEOPATRA Trastuzumab + PFS OS
cancer (Swain et al. 2015) docetaxel ± 18.7 versus 56.5 versus
pertuzumab 12.4 months; p 40.8 months; p
< 0.0001 = 0.0002
Breast cancer Neo-adjuvant 2012 NEOSPHERE pCR DFS
(Gianni et al. 2012) T+D 29% 81%
P+T+D 46% 86%
P+T 17% 73%
P+D 24% 73%
Trastuzumab Metastatic breast 2nd 2013 EMILIA (Verma T-DM1 versus ORR PFS OS
emtansine cancer et al. 2012) capecitabine + 44% versus 9.6 versus 6.4 30.9 versus
(T-DM1) lapatinib 31%; P < months; P < 25.1 months; p
0.001 0.001 < 0.001

(continued)
S. Parakh et al.
Table 1 (continued)
Target Drug Indication Tx Year Trial Treatment arms Endpoints
line
EGFR Cetuximab Metastatic 2004 Van Cutsem et al. Cetuximab + ORR PFS OS
colorectal (2009, 2011) FOLFIRI versus 57% versus 9.9 versus 8.4 23.5 versus 20
carcinoma FOLFIRI 40%; p < months; P = months; P =
0.001 0.0012 0.0093
Locally advanced 1st 2006 Bonner et al. Radiotherapy ± PFS OS
SCCHN (2006, 2010) cetuximab 17.1 versus 12.4 49 versus 29.3
months; p = 0.005 months; p =
0.018
Metastatic SCCHN 2nd 2011 EXTREME Platinum agent + ORR PFS OS
(Vermorken et al. fluorouracil ± 36% versus 5.6 versus 3.3 10.1 versus 7.4
2008) cetuximab 20%; p < months; P < months; P = 0.04
0.001 0.001
Panitumumab Metastatic 2nd 2006 Van Cutsem et al. Panitumumab versus PFS OS
KRAS WT (2007a) BSC 13.8 versus 8.5 No difference
colorectal cancer wks; P < 0.0001
Metastatic 1st 2006 PRIME (Douillard FOLFOX4 ± ORR PFS OS
colorectal cancer et al. 2010) panitumumab 54% versus 9.6 versus 8.0 23.9 versus 19.7
47% months, p = 0.02 months; P = 0.17
Current Development of Monoclonal Antibodies in Cancer Therapy

Necitumumab Metastatic SCC 1st 2015 SQUIRE Gemcitabine + ORR PFS OS

NSCLC (Paz-Ares et al. cisplatin ± 31.2% versus 5.7 versus 5.5 11.5 versus 9.9
2016) necitumumab 28.8%; p = 0.4 months; P = months; p =
0.006 0.012

(continued)
5
 6

Table 1 (continued)
Target Drug Indication Tx Year Trial Treatment arms Endpoints
line
VEGF Bevacizumab Metastatic 1st 2004 AVF2107 (Hurwitz FOLFIRI ± ORR PFS OS
colorectal et al. 2004) bevacizumab 45% versus 10.6 versus 6.2 20.3 versus 15.6
cancer 35% months; p < 0.001 months; p < 0.001
Metastatic 2nd 2006 ECOG E3200 ORR PFS OS
colorectal (Giantonio et al. FOLFOX versus 8.6% 4.7 months 10.9 months
cancer 2007)
FOLFOX + 22.7% 7.3 months 12.8 months
bevacizumab versus
Bevacizumab 3.3% (p < 2.7 (p < 0.001) 10.2 (P = 0.0011)
0.001)
Metastatic 1st 2006 ECOG E4599 Paclitaxel + ORR PFS OS
NSCLC (Sandler et al. 2006) carboplatin ± 35% versus 6.2 versus 4.5 12.3 versus 10.3
bevacizumab 15%; p < months; p < 0.001 months; p = 0.003
0.001
Metastatic 1st 2009 AVOREN (Escudier Interferon ± PFS OS
clear cell RCC et al. 2010) bevacizumab 10.2 versus 5.4 23.3 versus 21.3
months; p < months; p = .3360
0.0001
1st CALGB 90206 (Rini Interferon ± ORR PFS OS
et al. 2008, 2010) bevacizumab 25.5% versus 8.5 versus 5.2 18.3 versus 17.4
13.1% months; p < months; p = 0.097
0.0001
Glioblastoma 1st 2009 AVAGLIO (Chinot Temozolomide + RT PFS OS
multiforme et al. 2014) ± bevacizumab 10.6 versus 6.2 16.8 versus 16.7
months; p < 0.001 months; p = 0.10
2nd BELOB (Taal et al. OS
2014) Lomustine 43%
Bevacizumab 38%
Combination 63%

(continued)
S. Parakh et al.
Table 1 (continued)
Target Drug Indication Tx line Year Trial Treatment arms Endpoints
Metastatic EOC, 2nd 2014 AURELIA ICC ± ORR PFS OS
fallopian tube, or Platinum (Pujade-Lauraine chemotherapy 48% versus 8.2 versus 5.9 17 versus 13.3
peritoneal Ca resistant et al. 2014) 36%; p = months; p = months; p =
0.008 0.002 0.004
Metastatic EOC, 2nd 2014 OCEANS Gemcitabine + ORR PFS OS
fallopian tube, or Platinum (Aghajanian et al. carboplatin ± 79% versus 12.4 versus 8.4 33.6 versus
peritoneal Ca sensitive 2012, 2015) bevacizumab 57%; p < months; p < 32.9 months; p
0.0001 0.0001 = 0.65
Ramucirumab Metastatic gastric or 2nd line 2014 REGARD (Taal Ramucirumab PFS OS
GEJ adenocarcinoma et al. 2014) versus BSC 2.1 versus 1.3 5.2 versus 3.8
months; p < months; p =
0.001 0.047
Metastatic gastric or 2nd line 2014 RAINBOW Ramucirumab + ORR PFS OS
GEJ adenocarcinoma (Aghajanian et al. paclitaxel versus 28% versus 4.4 versus 2.9 9.6 versus 7.4
2012) paclitaxel 16%; p < months; p < months; p =
0.001 0.001 0.017
Metastatic NSCLC 2nd 2014 REVEL study Ramucirumab + PFS OS
(Garon et al. 2014) docetaxel versus 4.5 versus 3.0 10.5 versus 9.1
docetaxel months; p < months; p =
Current Development of Monoclonal Antibodies in Cancer Therapy

0.001 0.024
Metastatic colorectal 2nd 2015 RAISE (Horning FOLFIRI + PFS OS
cancer et al. 2005) ramucirumab 5.7 versus 4.5 13.3 versus
versus FOLFIRI months; p < 11.7 months p
0.001 = 0.023

(continued)
7
 8
Table 1 (continued)
Target Drug Indication Tx Year Trial Treatment arms Endpoints
line
PD1 Nivolumab Metastatic 2nd 2014 CheckMate 037 (Larkin et al. Nivolumab ORR PFS OS
melanoma 2016; Fuchs et al. 2014) versus ICC 27% versus 3.1 versus 3.7 16 versus 14
10% months months
Metastatic 2nd 2015 CheckMate 017 (Garon et al. Nivolumab ORR PFS OS
NSCLC 2014) versus docetaxel 20% versus 3.5 versus 2.8 9.2 versus 6.0
9% months; p = months; p =
0.31 0.0015
Metastatic RCC 2nd 2015 CheckMate 025 Nivolumab ORR PFS OS
versus 25% versus 4.6 versus 4.4 25.0 versus 19.6
everolimus 5%; p < months; p = months; p =
0.001 0.11 0.002
Metastatic 1st 2015 CheckMate 067 (Larkin et al. ORR PFS OS
melanoma 2016 2015; Wolchok et al. 2016) Nivolumab + 58.9% 11.7 months NR
ipilimumab
versus
Ipilimumab 44.6% 2.9 months 20 months
versus
Nivolumab 19% 6.9 months NR
Metastatic head 2nd 2016 CheckMate 141 (Ferris et al. Nivolumab PFS OS
and neck cancer 2016) versus ICC 2.0 versus 2.3 7.5 versus 5.1
months; p = months; p = 0.01
0.32
Metastatic 2nd 2017 CheckMate 275 (Sugimoto Nivolumab ORR OS
urothelial et al. 2003) 19.6% 7 months
cancer
Pembrolizumab Metastatic 1st 2014 KEYNOTE-006 (Robert ORR PFS OS
melanoma et al. 2015b; Long et al. Pembrolizumab 34% 5.5 months NR
2016) 10 mg/kg 2wkly
Pembrolizumab 33% 4.1 months NR
10 mg/kg wkly
Ipilimumab x4 3 12% 2.8 months NR
S. Parakh et al.

mg/kg q3wkly
(continued)
Table 1 (continued)
Target Drug Indication Tx Year Trial Treatment arms Endpoints
line
Metastatic 2nd 2015 KEYNOTE-010 (Ferrara 2010) PFS OS
NSCLC Pembrolizumab 3.9 months 10.4 months
2 mg/kg versus
Pembrolizumab 4.0 months 12.7 months
10 mg/kg versus
Docetaxel 4.0 months 8.5 months
Metastatic 1st 2016 KEYNOTE-024 (Reck et al. 2016) Pembrolizumab ORR PFS OS
NSCLC versus ICC 44.8% 10.3 versus 6.0 80.2% versus
versus months; p < 72.4%; p =
27.8% 0.001 0.005
Metastatic head 2nd 2016 KEYNOTE-012 (Teicher and Ellis Pembrolizumab ORR
and neck 2008; Hillen and Griffioen 2007) 18% (82% were durable responses  6 months)
cancer
PDL1 Avelumab Metastatic 2nd 2017 JAVELIN Merkel 200 (Telang Avelumab ORR
Merkel cell et al. 2011) 32% (86% were durable responses  6 months)
cancer
Atezolizumab Metastatic 2nd 2016 Rosenberg (Gerena-Lewis et al. Atezolizumab ORR
urothelial Ca 2009) 14.8% (84% were durable responses  6 months)
Metastatic 2nd 2016 OAK (Rittmeyer et al. 2017; Atezolizumab OS
NSCLC Barlesi et al. 2016) versus docetaxel 13.8 versus 9.6 months
Current Development of Monoclonal Antibodies in Cancer Therapy

CTLA-4 Ipilimumab Metastatic 2nd 2011 MDX010-20 (Hodi et al. 2010b) ORR OS
melanoma Ipilimumab + 10.9% 10 months
gp100 versus
Ipilimumab 5.7% 10 months
versus
gp100 alone 1.5% 6 months
BSC—Best supportive care; D—docetaxel; DFS—disease-free survival; FOLFOX—fluorouracil plus leucovorin and oxaliplatin; P—pertuzumab; TAC—paclitaxel, doxorubicin,
cyclophosphamide; T—trastuzumab; TTP—time to progression; ORR—overall response rates; OS—overall survival; CLEOPATRA—clinical evaluation of pertuzumab and trastuzumab;
ToGA—trastuzumab for gastric cancer; pCR—pathological complete response; RT—radiotherapy; SCC—squamous cell carcinoma
9
 10
Table 2 Approved monoclonal antibodies in haematological tumours
Target Drug Year Indication Study Key endpoints
CD20 Rituximab 2006 First-line treatment of DLBCL in
combination with CHOP or other
anthracycline-based chemotherapy
regimens
2006 First-line treatment of FL combined CVP + rituximab versus CVP (Marcus et al. ORR TTP TTnT
with CVP and following CVP 2005) 81% versus 32 versus 27 versus
57% 15 months 7 months
(p < 0.0001) (p < 0.0001) (p < 0.0001)
2010 In combination with FC for the Rituximab ± FC (Hallek et al. 2010; Fischer et al. ORR PFS OS
treatment of CLL in untreated and 2012) 86% versus 42.5 versus NR versus
previously treated patients 73% 33.1 months 86.0 months
(p = 0.02) (p = 0.001)
Rituximab ± FC (Robak et al. 2010) ORR PFS OS
61% versus 27 versus NR versus
49% 21.9 months 52 months
(p < 0.02) (p = 0.2874)
2011 Maintenance therapy in untreated Rituximab maintenance versus observation (Salles PFS No
FL et al. 2011) 74.9% versus 57.6% difference in
(p < 0.0001) OS
90
Y 2002 Relapsed or refractory, low-grade Single-arm phase II (Wiseman et al. 2002) ORR TTP DoR
Ibritumomab follicular B cell NHL or 83% 9.4 months 11.7 months
tiuxetan rituximab-refractory follicular NHL
90
2009 Treatment of previously untreated Y Ibritumomab tiuxetan versus no treatment PFS TTnT
follicular NHL, who achieve an (Hagenbeek et al. 2007; Morschhauser et al. 4.1 versus 8.1 versus
objective response to first-line 2013) 1.1 years 3 years
chemotherapy (p < 0.001) (p < 0.001)
131
I 2003 FL refractory to rituximab and Single-arm phase II (Horning et al. 2005) ORR TTP DoR
Tositumomaba relapsed following chemotherapy 63% 10.4 months 16 months
Ofatumumab 2014b In combination with chlorambucil, Chlorambucil + ofatumumab versus ofatumumab PFS
for previously untreated CLL where (Hillmen et al. 2015) 22.4 versus 13.1 months (p < 0.001)
fludarabine-based therapy is
S. Parakh et al.

considered inappropriate
(continued)
Table 2 (continued)
Target Drug Year Indication Study Key endpoints
2016 In combination with FC in relapsed FC + ofatumumab versus FC (Robak et al. 2017) ORR DoR PFS
CLL 84% versus 29.6 versus 28.9 versus
68% 24.9 months 18.8 months
(p = 0.0878) (p = 0.0032)
Obinutuzumab 2013 In combination with chlorambucil Obinutuzumab + chlorambucil versus ORR DoR PFS
for previously untreated CLL chlorambucil (Goede et al. 2014, 2015) 75.9% 15.2 versus 23 versus
versus 3.5 months 11.1 months
32.1% (p < 0.001)
2016 In combination with bendamustine Obinutuzumab + bendamustine -> obinutuzumab ORR DoR PFS
followed by obinutuzumab monotherapy versus bendamustine 78.7% NR versus NR versus
monotherapy for patients with FL versus 11.6 months 13.8 months
refractory to a rituximab (Sehn et al. 74.7% (p < 0.
2016) 0001)
CD30 Brentuximab 2011 Hodgkin lymphoma after ASCT or Single-arm phase II studies (Younes et al. 2012; ORR PFS OS
vedotin  2 prior chemotherapy regimens in Gopal et al. 2014) 75% 40.5 months 9.3 months
patients not candidates for ASCT
2011 Systemic ALCL after prior Single-arm study (de Claro et al. 2012) ORR DoR
multi-agent chemotherapy regimen 86% 12.6 months
2015 Patients with HL at high risk of Brentuximab vedotin versus placebo PFS
relapse/progression post-ASCT 42.9 versus 24.1 months (p = 0.0013)
consolidation
Current Development of Monoclonal Antibodies in Cancer Therapy

CD38 Daratumumab 2016 For treatment of relapsed MM in Daratumumab ± lenalidomide + dexamethasone ORR DoR PFS
combination with lenalidomide or (Dimopoulos et al. 2016) 93% versus NR versus NR versus
bortezomib and dexamethasone 76% 17.4 months 18.4 months
(p < 0.0001) (p < 0.0001)
Daratumumab ± bortezomib + dexamethasone ORR TPP PFS
(Palumbo et al. 2016) 83% versus NR versus NR versus
63% 7.3 months 7.2 months
(p < 0.0001) (p < 0.0001) (p < 0.0001)
(continued)
11
 12
Table 2 (continued)
Target Drug Year Indication Study Key endpoints
PD-1 Nivolumab 2016 HL relapsed or progressed after Single-arm studies (Timmerman et al. 2016; ORR DoR
auto-HSCT and post-transplantation Ansell et al. 2015) 65% 8.7 months
brentuximab vedotin
Pembrolizumab 2017 Refractory classical Hodgkin Single-arm study (Chen et al. 2016b; Moskowitz ORR DoR PFS
lymphoma et al. 2016) 69% NR (6 months)
72.4%
SLAMF7 Elotuzumab 2015 In combination with lenalidomide Elotuzumab ± lenalidomide and dexamethasone ORR PFS
and dexamethasone for multiple 75.8% 19.4 versus
myeloma who received  1 versus 14.9 months
therapies 65.5%
ASCT—Autologous stem cell transplantation; ALCL—anaplastic large cell lymphoma; CR—complete response; CVP—cyclophosphamide, vincristine and prednisone; CHOP
—cyclophosphamide, doxorubicin, vincristine and prednisone; CLL—chronic lymphocytic leukaemia; DoR—duration of response; HL—Hodgkin’s lymphoma; HSCT—
haematopoietic stem cell transplantation; FC—fludarabine and cyclophosphamide; FL—follicular lymphoma; NHL—non-Hodgkin’s lymphoma; NR—not reached; ORR—
overall response rate; PFS—progression-free survival; TTP—time to progression; TTnT—time to next treatment
a
Discontinued due to projected decline in sales and the availability of alternative treatments
b
Initial approval in 2009 as part of the FDA’s accelerated approval process
S. Parakh et al.
Current Development of Monoclonal Antibodies in Cancer Therapy 13

Target antigens are not restricted to the tumour itself, as tumour support struc-
tures like the vasculature, stroma and extracellular matrix also provide potential
targets (Scott et al. 2012; Ferris et al. 2010). Vascular endothelial growth factor
(VEGF) is one of the ligands for the VEGF receptor 2 (VEGFR2) where it plays a
role in promoting angiogenesis in developing tumours, and both have been targeted
by antibodies in cancer therapy (Holmes et al. 2007). Trapping of the ligand or
blocking of the receptor is able to limit the tumours’ ability to develop vascular
networks, thereby reducing its capacity to grow and spread (Holmes et al. 2007).
Additional to tumour support structures, the immune system itself can be targeted in
cancer therapy to enhance the natural response against the tumour. For example,
cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) is a cell surface receptor
expressed on T cells that when activated by its ligand it functions to downregulate
their responses (Leach et al. 1996; Hodi et al. 2010a). Ipilimumab is an antibody
that has been developed to block CTLA-4 cytotoxic T cells, and this blocking of
CTLA-4 keeps the T cells in an active state with anti-tumour capacity (Hodi et al.
2010a).

1.2 Monoclonal Antibody Formats

Monoclonal antibodies (mAbs) are produced by a single B cell clone and target one
specific epitope of an antigen. The first method for the production of mAbs was the
hybridoma technology introduced by Kohler and Milstein in 1975 (Kohler and
Milstein 1975). This method involves immunizing mice with the antigen of interest
and then isolating B cells from the spleen. These isolated B cells are then fused with
myeloma cells with each fusion resulting in an immortal cell that produces
unlimited quantities of identical antibody, called hybridomas. The first mAb
approved for cancer therapy, rituximab, is used for the treatment of non-Hodgkin’s
lymphoma through targeting of the CD20 receptor on B cells (Maloney et al. 1997).
Even though the technology existed to produce large quantities of antibodies to
any target antigen required, there were still several obstacles to overcome in order
to achieve maximal efficacy. Because hybridoma-derived antibodies were of murine
origin, their therapeutic potential is hampered by two main problems. Firstly, the
murine Fc region of the antibodies has reduced binding to human Fc receptors
which impair cellular effector function of immune cells and diminishes serum
half-life. Secondly and most importantly, the infusion of murine antibodies into
patients leads to the development of a host immune response to the foreign protein
and the production of human anti-mouse antibodies (HAMA).
To overcome HAMA responses and improve the interaction with human Fc
receptors, methods to produce antibodies with higher human homology were
established. The first methods developed took advantage of recombinant DNA
technology by isolating the mRNA sequence coding for the antibody from the
hybridoma. With the murine DNA, it was then possible to substitute in human
constant region DNA to reduce immunogenicity without impacting on antigen
binding. This combination produced chimeric antibodies with murine variable
14 S. Parakh et al.

regions and human constant regions (Morrison 1985; Neuberger et al. 1985;
Norderhaug et al. 1997). Following the success of chimeric antibodies, further
efforts to produce mAbs with higher human homology were researched. These
chimeric antibodies could be refined further by selective alteration of the amino
acids in the framework region of the variable domain portion while still keeping the
original murine CDRs (Riechmann et al. 1988; Queen et al. 1989). This process of
refinement is referred to as humanization and produces mAbs where the only
murine sequence is limited to the CDRs.
With the technology established to produce chimeric and humanized mAbs,
research continued to design methods to create “fully” human mAbs (Laffleur et al.
2012). Following the creation of transgenic mice with human germline genes for
the heavy and kappa light chains, it was possible to immunize these mice to
produce human antibodies that were antigen-specific (Lonberg and Huszar 1995).
Platforms to produce mAbs without the use of mice were also developed that use
phage display to randomly generate CDRs in Fab fragments (Winter and Milstein
1991; Smith 1985; Bazan et al. 2012). This is achieved through the use of viruses
that infect bacteria (bacteriophages) that express proteins on the viral particle sur-
face, and DNA for these random CDRs is inserted into genes for surface proteins
(Bazan et al. 2012). In doing this, a large library of phage surfaces can be generated
and screened against the antigen of interest, thereby creating a high-throughput
method for generating novel antibodies (Bazan et al. 2012). Once identified, any
positive clones can also be further refined to enhance binding through small amino
acid changes in the CDRs through iterations of the phage display system (Bazan
et al. 2012).
Additional to the use of full-sized antibodies, it is possible to produce smaller
antibody fragments that retain antigen-binding activity. Due to their smaller size,
these antibody fragments have altered pharmacokinetic properties that make them
useful for the diagnosis and treatment of cancers (Chames et al. 2009; Holliger and
Hudson 2005). These include monovalent Fab fragments, approximately 55 kDa,
which lack an Fc and are not able to engage effector functions through FcRs.
Without FcR engagement, Fab fragments have a short half-life, 12–20 h, compared
to intact IgG molecules which have a half-life in serum of more than 10 days
(Holliger and Hudson 2005; Flanagan and Jones 2004). This short half-life of Fab
fragments makes them great tools for imaging tumours because of the rapid blood
clearance. Arcitumomab is a Fab fragment of a murine monoclonal antibody that
targets carcinoembryonic antigen expressed on colorectal cancers and has been used
as a radioimmunoconjugate with the radioisotope technetium-99 m for tumour
imaging (Hansen et al. 1990). As a result of the rapid blood clearance of arcitu-
momab the background observed when imaging tumours is reduced when compared
to the intact IgG (Behr et al. 1995). Fab fragments are not the only antibody formats
available, and single-chain variable fragments (scFv) are created by linking both the
heavy and light variable domains of an antibody with flexible polypeptide linker
(Holliger and Hudson 2005). This creates an even smaller molecule, approximately
28 kDa, that still retains antigen-binding capacity, and multiple scFvs can be linked
together to create multivalent complexes (Holliger and Hudson 2005).
Current Development of Monoclonal Antibodies in Cancer Therapy 15

1.3 Mechanism of Action

Antibodies used in cancer therapy have various methods of mediating tumour cell
death, which are intimately linked with the native function of the target antigen
(Fig. 2). They can directly act on tumour cells by the blocking of growth factor
receptors that are required for tumour growth (Scott et al. 2012) such as members of
the ErbB family. Cetuximab is one such example, and it was the first monoclonal
antibody to target EGFR and has been approved for the treatment of colorectal
cancer patients (Van Cutsem et al. 2009). By binding to EGFR on the cell surface,
cetuximab blocks the ligand binding site which in turn inhibits intracellular receptor
signalling resulting in cell-cycle arrest, induction of apoptosis and downregulation
of cell surface expression of EGFR. Trastuzumab is another example of a mAb that
has been successfully used to target the human epidermal growth factor receptor
type 2 (HER2).
Antibodies can also act upon stromal cells and vasculature in the tumour
microenvironment to limit growth or induce tumour cell death (Scott et al. 2012).
Malignant tumours are made up of rapidly dividing and growing cancer cells, and in
order to support this growth, they need their own dedicated blood supply. This is
achieved by the tumour releasing factors that tilt the balance within the microen-
vironment towards pro-angiogenesis to drive vascular growth and establish its own
blood network. Bevacizumab is an antibody that targets the pro-angiogenic factor,
vascular endothelial growth factor A (VEGF-A) (Yang et al. 2003). Bevacizumab
works by binding to and trapping the soluble form of VEGF-A, which stops
VEGF-A from working as a ligand to stimulate the vascular endothelial growth
factor receptor (VEGF) expressed on endothelial cells (Willett et al. 2004).
Antibody-coated tumour cells are recognized by immune cells via interactions
with specific receptors on their cell surface and trigger cellular effector functions.
This interaction is mediated by the Fc domain of the antibody which contains
specific binding sites for receptors on the immune cell surface (Hogarth and Pietersz
2012; Nimmerjahn and Ravetch 2008). These are known as the Fc receptors (FcRs),
which are expressed on a variety of immune effector cells. Natural killer (NK) cells
have the greatest reputation among anti-tumour effector cells as they have been
shown to be the primary mediators of antigen-dependent cell-mediated cytotoxicity
(ADCC) (Hogarth and Pietersz 2012; Nimmerjahn and Ravetch 2008). ADCC is
triggered when the Fc domain of the bound therapeutic antibody is recognized by
Fc gamma receptor IIIa (FccRIIIa) on the surface of NK cells and involves the
release of cytotoxic factors, such as perforin, that cause lysis of the tumour cell
(Hogarth and Pietersz 2012; Nimmerjahn and Ravetch 2008). ADCC can be
enhanced in therapeutic antibodies through engineering that improves the interac-
tion of the antibody Fc domain with FccRIIIa (Desjarlais and Lazar 2011; Des-
jarlais et al. 2007). This can be achieved through amino acid engineering, whereby
a more favourable amino acid can be introduced to the enhance interaction with Fc
(Desjarlais and Lazar 2011; Desjarlais et al. 2007). Additionally, the alteration of
the glycosylation pattern of IgG Fc to reduce fucose content can provide a selective
binding enhancement to FccRIIIa and improved ADCC (50–100-fold increase).
16 S. Parakh et al.

Fig. 2 Mechanisms of action of monoclonal antibodies: a Immune-mediated tumour cell

killing through complement activation, antibody-dependent cellular cytotoxicity (ADCC),
inhibition of T cell inhibitory receptors such as cytotoxic T lymphocyte-associated antigen 4
(CTLA4) and induction of phagocytosis; b tumour cell killing through inhibition of dimerization,
kinase activation and downstream signalling leading to reduced proliferation and apoptosis;
c conjugated antibodies to deliver toxic payloads such as a drug, toxin, small interfering RNA or
radioisotope into tumour-cell-inducing cell death

Opsonized tumour cells can be phagocytosed through engagement of FccRs, and

in addition to the killing of the target cell, antigen presentation can occur to activate
T cells (Nimmerjahn and Ravetch 2008; Richards et al. 2008). The process of
engulfing an antibody-coated tumour cell is called antigen-dependent cellular
phagocytosis (ADCP) and involves the engagement of activatory FccRs (Nim-
merjahn and Ravetch 2008; Richards et al. 2008). This could potentially provide
patients with long-term immunity against tumours through the induction of the
adaptive immune system in the form of anti-tumour memory T cells (Richards et al.
2008). This linking of the innate and adaptive immune system through the passive
administration of anti-tumour antibodies is referred to as the vaccinal effect and has
long been thought as the holy grail of antibody therapy (Richards et al. 2008;
Current Development of Monoclonal Antibodies in Cancer Therapy 17

DiLillo and Ravetch 2015). An additional Fc-mediated function of antibodies is to

engage the complement system through a number of small proteins found in the
blood (Duncan and Winter 1988; Ricklin et al. 2010). This involves the recruitment
of these proteins to form a complex on the surface of the pathogen (Duncan and
Winter 1988; Ricklin et al. 2010). This complex serves three main functions: to
enhance phagocytosis, recruit immune cells, and form a membrane attack complex
(MAC) that lyses the target (Ricklin et al. 2010).
Antibodies can also be used to deliver a payload directly to the tumour site due
to the unique specificity for their TAA. These antibody conjugates can be used to
deliver a drug, toxin, small interfering RNA (siRNA) or even a radioactive isotope
in a targeted approach that often leads to improved efficacy and reduced toxicity.
Trastuzumab is used as an antibody–drug conjugate (ADC) to deliver the
chemotherapeutic agent DM1 directly to the tumour through receptor internaliza-
tion (Barok et al. 2014; Hudis 2007). The use of trastuzumab as an ADC allows for
a broad anti-tumour mechanism of action with its ability to abrogate HER2 sig-
nalling, recruit immune cells and directly kill the tumour through payload delivery.

2 Approved Monoclonal Antibodies

A summary of approved monoclonal antibodies in cancer treatment is detailed in

Tables 1 and 2 with an approval history timeline shown in Fig. 3.

2.1 Anti-CD20 Monoclonal Antibodies

CD20 is a transmembrane calcium channel highly expressed on the surface of

human B cells, making it an ideal target for directed therapy. It is involved in B cell
activation, proliferation, differentiation (Gopal and Press 1999) and calcium flux
(Bubien et al. 1993). Cross-linking of CD20 with monoclonal antibodies has shown
to trigger antibody-dependent cellular cytotoxicity (ADCC) and complement-
dependent cytotoxicity (CDC) in cells (Kosmas et al. 2002).

2.1.1 Rituximab
Rituximab is a chimeric anti-CD20 monoclonal antibody, which has revolutionized
the management of B cell lymphoproliferative malignancies. Rituximab is one of
the most widely prescribed biological agents today. It was first approved in 1997 for
the treatment of relapsed indolent NHL. Today, it is used in a number of settings as
induction therapy, for maintenance of disease remission and at disease relapses in
NHL as well as in chronic lymphocytic leukaemia (CLL).
In 2006, rituximab was approved for use as first-line treatment in patients with
diffuse large B cell (DLBCL) in combination with cyclophosphamide, doxorubicin,
vincristine and prednisone (CHOP) or other anthracycline-based chemotherapy
regimens based on the results of three randomized trials involving nearly 2000
treatment-naive patients (Pfreundschuh et al. 2010; Coiffier et al. 2010; Habermann
18 S. Parakh et al.

Fig. 3 Timeline of antibody approval in cancer treatment

et al. 2006). Two trials evaluated patients aged  60 years with stage 3–4 disease
(E4494 (Habermann et al. 2006) and LNH 98-5/GELA (Coiffier et al. 2010)), while
the M39045/MiNT (Pfreundschuh et al. 2010)-enrolled patients aged between 18
and 60 years with majority having early-stage disease. In each study, hazard ratios
for the main outcome measured as well as overall survival favoured the
rituximab-containing arms, with results consistent across all subgroup analysis. The
benefit of rituximab was seen at long-term follow-up (Feugier et al. 2005).
The benefit of rituximab in this setting however has shown to be limited to patients
whose tumours do not express the bcl-6 protein (Winter et al. 2006). Also in 2006,
rituximab was approved for use as first-line treatment for patients with low-grade or
follicular B cell, CD20-positive NHL, in combination with cyclophosphamide,
vincristine and prednisone (CVP) as well as following CVP chemotherapy (Marcus
et al. 2005). Treatment with single-agent rituximab after CVP chemotherapy in
patients that have responded resulted in a statistically significant reduction in PFS.
In 2010, rituximab in combination with fludarabine and cyclophosphamide
(FC) was approved for the treatment of CLL in treatment-naive patients with
excellent performance status as well as in patients with relapsed or refractory
disease based on the positive findings of two randomized trials, ML17102 and
BO17072, respectively (Hallek et al. 2010; Robak et al. 2010; Fischer et al. 2012).
Current Development of Monoclonal Antibodies in Cancer Therapy 19

The benefit of FCR however was not seen in a subgroup of patients with del(17p)
by fluorescence in situ hybridization (FISH)1, TP53 mutations (Rossi et al. 2014)
and unmutated immunoglobulin heavy chain variable (IGHV) gene (Thompson
et al. 2016).
Rituximab in 2011 was approved for maintenance therapy in patients with
previously untreated follicular CD20-positive B cell NHL after first-line treatment
with rituximab in combination with chemotherapy. The approval was based on
phase III PRIMA trial (Salles et al. 2011). Despite the benefit in PFS, this did not
translate into an improvement in overall survival or reduce the rate of histological
transformation. With no clear benefit in overall survival, reported in a number of
other randomized trials, there remains a lot of debate with regard to the optimum
duration of maintenance treatment.

2.1.2 Ofatumumab
Ofatumumab is a humanized type I anti-CD20 monoclonal antibody approved for
the treatment of CLL in treatment-naive patients as well as patients with relapsed,
treatment-refractory disease. A single-arm study evaluated ofatumumab in patients
with CLL refractory to fludarabine and alemtuzumab and in patients with
fludarabine-refractory CLL with bulky (>5 cm) lymphadenopathy who were not
suitable for alemtuzumab. Treatment with ofatumumab resulted in improved
responses rates and complete resolution of constitutional symptoms and improved
performance status in almost half of all patients (Wierda et al. 2010).
In a phase III trial (COMPLEMENT 1) (Hillmen et al. 2015), the addition of
ofatumumab to chemotherapy in treatment-naive patients with CLL resulted in a
significant improvement in median PFS and prolonged median duration of
response. After a median follow-up of 28.9 months, OS was not reached in both
groups. Despite enrolling elderly patients (half of enrolled patients >70 years) and
those with multiple comorbidities, the addition of ofatumumab resulted in clinically
meaningful improvements.
In the COMPLEMENT 2 trial (Robak et al. 2017), patients with relapsed CLL
were randomized to fludarabine and cyclophosphamide with or without ofatu-
mumab. The addition of ofatumumab resulted in significantly longer PFS, with
manageable toxicities.

2.1.3 Obinutuzumab
Obinutuzumab is the first Fc-engineered, humanized anti-CD20 monoclonal anti-
body to be approved in combination with chlorambucil in the management of
treatment-naive patients with CLL. In the randomized phase III trial, CLL11, the
median age of patients was 73 years, 68% had impaired renal function, and 76%
had multiple coexisting medical conditions. Chemo-immunotherapy with either
rituximab or obinutuzumab was shown to be superior to chemotherapy alone. After
a follow-up of 39 months, compared with rituximab, treatment with obinutuzumab
resulted in clinically meaningful PFS and TTNT with a trend towards improved OS
20 S. Parakh et al.

(Goede et al. 2014, 2015). In a subgroup analysis defined by gene mutations,

obinutuzumab–chlorambucil had better outcomes and was able to overcome
NOTCH1mut-associated rituximab resistance (Estenfelder et al. 2016).
In 2016, obinutuzumab was approved in combination with bendamustine fol-
lowed by obinutuzumab monotherapy as maintenance therapy for the treatment
rituximab-refractory follicular lymphoma based on the findings of the GADOLIN
study, in which the median PFS was significantly higher in patients treated with
obinutuzumab plus bendamustine (Sehn et al. 2016).
Toxicities Associated with Anti-CD20 Antibodies
Infusion reactions are the most commonly reported adverse event associated with
anti-CD20 antibodies (Coiffier et al. 2008; Ghielmini et al. 2005; Kasi et al. 2012).
Infusion reactions typically occur after the first infusion and within 24 h of the
infusion and are dose-dependent (Ghielmini et al. 2005; Coiffier et al. 1998). Most
infusion reactions are mild with severe (grade 3/4) reactions rare (Kasi et al. 2012).
Grade 3/4 cytopenias have been reported in almost half of all patients treated with
anti-CD20 antibodies with lymphopenia and neutropenia being the most commonly
reported (Coiffier et al. 2008; Kasi et al. 2012). This incidence increases when
rituximab is used in combination with chemotherapy regimens (Buske et al. 2009;
Eve et al. 2009). There is an associated increase in frequency of infectious com-
plications, with bacterial infections most commonly seen (Kasi et al. 2012; Cohen
et al. 2006). A dose-dependent increase in the frequency of infections is seen in
patients treated with rituximab (Avilés et al. 2007). Severe and sometimes fatal
mucocutaneous reactions can occur. Also reported is JC virus reactivation, leading
to progressive multifocal leukoencephalopathy (Carson et al. 2009). Pulmonary
complications have been reported in 5% of patients treated with rituximab
monotherapy. Hypoalbuminemia has been identified as an independent risk factor
(Kang et al. 2012). The most common toxicities are infectious; however, other
toxicities reported include interstitial lung disease, bronchiolitis obliterans, hyper-
sensitivity pneumonitis and diffuse alveolar haemorrhage (Biehn et al. 2006;
Tonelli et al. 2009; Heresi et al. 2008).
Resistance Mechanisms to Anti-CD20 Antibodies
The exact mechanisms of rituximab resistance remain poorly understood. Resis-
tance mechanisms can be broadly divided into host factors such as Fc receptor
polymorphisms affecting the affinity of effector cells for rituximab Fc and
tumour-related factors, e.g. CD20 expression and structure, acquired CD20 muta-
tions and tumour burden. Other general mechanisms include alterations in ritux-
imab pharmacokinetics. Rituximab-resistant cell lines have shown to express high
levels of membrane complement regulatory proteins, CD20, CD55 and CD59,
which inhibit the complement cascade, thereby affecting the ability of rituximab to
induce complement-dependent cytotoxicity (Takei et al. 2006). Patients with
rituximab resistance transfused with fresh frozen plasma and treated with rituximab
showed excellent clinical response (Klepfish et al. 2009; Xu et al. 2011).
Current Development of Monoclonal Antibodies in Cancer Therapy 21

The efficacy of ofatumumab is limited by drug resistance, which is not well

characterized (Baig et al. 2010). In vitro studies using CLL cells pre- and
post-treatment with ofatumumab showed a single dose of ofatumumab resulted in a
marked decrease in serum complement levels, and the surviving CLL cells showed
depletion in CD20 expression. These cells were resistant to in vitro ofatumumab-
mediated CDC, but most retain full sensitivity to alemtuzumab-mediated CDC.

2.2 Anti-CD30 Monoclonal Antibodies

CD30, a member of the tumour necrosis factor receptor (TNFR) family, is a

transmembrane glycoprotein with expression in normal tissues limited to some
activated B and T cells (Younes and Kadin 2003). CD30 is highly expressed in
Hodgkin lymphoma (HL) and systemic anaplastic large cell lymphoma (sALCL)
cells irrespective of disease stage, line of therapy or transplant status (Francisco
et al. 2003). In contrast, CD30 expression in solid tumours occurs at a lower
frequency compared to haematopoietic-derived tumours; however, high expression
of CD30 has shown to occur in testicular embryonal carcinoma and germ cell
tumours (Dürkop et al. 2000).

2.2.1 Brentuximab Vedotin

Brentuximab vedotin (BV) is an antibody–drug conjugate (ADC) composed of a
monoclonal antibody directed against CD30 that is covalently bound by a
protease-cleavable linker to the anti-microtubule agent monomethyl auristatin E
(MMAE) (Francisco et al. 2003). Binding of BV to CD30 results in internalization
of the MMAE-CD30 complex and release of MMAE by proteolytic cleavage.
In 2011, BV received accelerated approval for the treatment of patients with
CD30-positive HL that relapsed after autologous stem cell transplantation (ASCT),
and relapsed sALCL, based on results of two single-arm trials (de Claro et al. 2012;
Chen et al. 2016a). In 2015, brentuximab vedotin also received FDA approval for
the treatment of patients with HL at high risk of relapse/progression post-ASCT
consolidation. This approval was based on the AETHERA trial which showed an
improvement in the median PFS in the BV group compared to 24.1 months in the
placebo group (Moskowitz et al. 2015). This benefit was seen in all subgroups
analysed.
Toxicities
The most frequent adverse events related to BV seen across all trails were peripheral
sensory neuropathy, neutropenia, fatigue and nausea. Other toxicities reported were
diarrhoea, pyrexia, upper respiratory tract infection and vomiting. In all trials,
neuropathy was the leading cause of treatment discontinuation. Toxicity usually
developed between 3 and 4 months after commencing treatment (Scott 2017;
Garnock-Jones 2013). At the five-year follow-up of patients with relapsed/refractory
HL who developed treatment-related peripheral neuropathy, majority experienced
either resolution or improvement in symptoms (Chen et al. 2016a).
22 S. Parakh et al.

Mechanisms of Resistance
Currently, mechanisms of resistance to BV are unknown (Chen et al. 2015). Using
two different treatment models, Chen et al. (2015) developed BV-resistant HL
(L428) and ALCL (Karpas-299) cell lines to elucidate potential resistance mech-
anisms. Although loss of target expression was not shown in the BV-resistant HL
cell line or in tissue samples of patients with HL who had relapsed or progressed
after BV treatment, the alteration in signalling level may be a potential mechanism.
A reduction in the dynamics of receptor cellular or receptor internalization could
also reduce the efficiency of antigen targeting (Parakh et al. 2016a). In contrast, the
HL cell line, but not the ALCL cell line, exhibited MMAE resistance and over-
expression of MDR1 mRNA compared to the parental line. Both HL and ALCL
treatment-resistant patient samples persistently expressed CD30 by immunohisto-
chemistry (Chen et al. 2015).

2.3 Anti-CD38 Monoclonal Antibodies

CD38 is a transmembrane glycoprotein expressed by a variety of lymphoid and

myeloid lineages; in particular, plasma cells express particularly high levels of
CD38 (Deaglio et al. 2001). CD38 functions as an adhesion molecule, involved in
the activation and proliferation of human leucocytes as well as signal transduction
and intracellular calcium mobilization (Lin et al. 2004; Malavasi et al. 1994). CD38
is highly expressed in a number of haematological malignancies, in particular MM
(Lin et al. 2004) and CLL (Damle et al. 1999). CD38 expression by CLL cells has
shown to be associated with a more aggressive clinical course and poorer patient
outcomes (Damle et al. 1999).

2.3.1 Daratumumab
Daratumumab is a humanized anti-CD38-specific antibody. In 2016, daratumumab
was approved for the treatment of multiple myeloma (MM) in combination with
lenalidomide and dexamethasone, or bortezomib and dexamethasone, in patients
who have received prior therapy. The approval is based on two randomized trials in
which daratumumab in combination with standard therapies resulted in improved
response rates and PFS (Plesner et al. 2014; Palumbo et al. 2016; Dimopoulos et al.
2016).
Toxicities
The most frequently reported haematological AEs of any grade were anaemia,
thrombocytopenia and neutropenia. Nearly half of all patients experienced
infusion-related reactions post-cycle 1, majority of which were of low grade.
Common non-haematological AEs were fatigue, nausea and diarrhoea (Sanchez
et al. 2016). Another effect unique to anti-CD38 monoclonal antibodies is the high
false positive results with the indirect anti-globulin test (Coombs test) (Oostendorp
Current Development of Monoclonal Antibodies in Cancer Therapy 23

et al. 2015) as daratumumab binds to CD38 expressed on RBC, masking antigens in

the patient’s serum (Oostendorp et al. 2015).
Mechanisms of Resistance
Majority of patients treated with daratumumab eventually develop resistance
(Nijhof et al. 2016). Examining patient samples pre- and post-treatment with
daratumumab showed CD38 expression levels correlated with response, while
expression of complement-inhibitory proteins (CIPs), membrane cofactor protein
(CD46), decay-accelerating factor (CD55) and protectin (CD59) levels increased at
time of progression and correlated with development of antibody resistance.
Treating MM cells from patients who developed daratumumab resistance with
ATRA led to increased CD38 levels and decreased CD55 and CD59 expression to
almost pretreatment values (Nijhof et al. 2015, 2016). Furthermore, in the light of
the immune-effector-mediated mechanism of daratumumab, T cell exhaustion could
affect its effectiveness.

2.4 Anti-CD52 Monoclonal Antibodies

CD52 is a glycosylphosphatidylinositol-anchored antigen highly expressed on

normal and neoplastic lymphoid cells. Erythrocytes, platelets and bone marrow
stem cells lack CD52 surface expression (Xia et al. 1991). The CD52 antigen is also
expressed on subsets of tumour cells, including T cell prolymphocytic leukaemia,
CLL, hairy cell leukaemia, NHL and acute lymphoblastic leukaemia (Ginaldi et al.
1998). The exact biological function of CD52 is yet to be elucidated; however,
some evidence suggests that it may be involved in T cell migration and costimu-
lation (Watanabe et al. 2006).

2.4.1 Alemtuzumab
Alemtuzumab is an anti-CD52 humanized monoclonal antibody approved in 2007
for use as a single agent in the treatment of patients with B cell CLL (B-CLL)
refractory to alkylating agents and failed fludarabine therapy. Alemtuzumab ini-
tially received accelerated approval in 2001 based on the findings of three
single-arm phase II studies (Osterborg et al. 1997; Rai et al. 2002; Keating et al.
2002). Full approval was given following results of the larger randomized phase III
trial, CAM 307 (Hillmen et al. 2007).
Toxicities
The most common adverse events associated with alemtuzumab are infusion-related
side effects, myelosuppression and infections (Fraser et al. 2007). Grade 3/4 reac-
tions however were noted in up to 20% of patients. The incidence of
infusion-related side effects was similar regardless of the treatment setting and was
most severe on first exposure to the drug (Osterborg et al. 1997; Rai et al. 2002;
Keating et al. 2002; Liggett et al. 2005; Ferrajoli et al. 2003). The subcutaneous
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