Primary resistance to crizotinib treatment in a non-small cell lung cancer patient with an EML4-ALK rearrangement: a case report

Abstract

Crizotinib, a small molecular tyrosine kinase inhibitor, manifests dramatic responses in patients with non-small cell lung cancer with echinoderm microtubule associated protein like 4-anaplastic lymphoma kinase (EML4-ALK) rearrangements. ALK gene point mutation is the primary mechanism of acquired crizotinib resistance; however, the intrinsic mechanism is not fully understood. Here, we report a patient with a low mutant allele fraction (MAF) of EML4-ALK rearrangement, who experienced primary resistance to crizotinib treatment. The patient was a 66-year-old Chinese man, who had a history of metastatic lung cancer and was treated with first- and third-generation epidermal growth factor receptor tyrosine kinase inhibitors (EGFR TKIs). After 14 months of osimertinib treatment, his disease progressed, and next-generation sequencing was performed from a liquid biopsy of the patient's blood. An EML4-ALK rearrangement was found and crizotinib was administered. The patient's lung lesions continued to progress after one month of crizotinib treatment, and pemetrexed-bevacizumab was initiated. After two cycles of chemotherapy, the metastatic cancers shrunk, and the patient maintained stable disease at his last follow-up. EML4-ALK rearrangements can happen in patients with EGFR-positive NSCLC, after acquired resistance to EGFR TKI treatment. The EGFR T790M and C797G mutations occur in cis is a critical mechanism of resistance to osimertinib therapy. The MAF of EML4-ALK rearrangements in cancer cells might be a predictive factor for crizotinib treatment.

Full text

CASE REPORT
Primary resistance to crizotinib treatment in a non-small cell
lung cancer patient with an EML4-ALK rearrangement: a case
report
Ling Zhang1, Yunxia Li1, Shaohong Zhang2, Chen Gao1, Keke Nie1, Youxin Ji3
1Department of Oncology, Qingdao Cancer Hospital, Qingdao 266042, China; 2Department of Medicine, Laixi Meihuashan
Hospital, Qingdao 266600, China; 3Department of Oncology, Affiliated Qingdao Central Hospital of Qingdao University,
Qingdao 266042, China

ABSTRACT Crizotinib, a small molecular tyrosine kinase inhibitor, manifests dramatic responses in patients with non-small cell lung cancer
with echinoderm microtubule associated protein like 4-anaplastic lymphoma kinase (EML4-ALK) rearrangements. ALK gene
point mutation is the primary mechanism of acquired crizotinib resistance; however, the intrinsic mechanism is not fully
understood. Here, we report a patient with a low mutant allele fraction (MAF) of EML4-ALK rearrangement, who experienced
primary resistance to crizotinib treatment. The patient was a 66-year-old Chinese man, who had a history of metastatic lung
cancer and was treated with first- and third-generation epidermal growth factor receptor tyrosine kinase inhibitors (EGFR TKIs).
After 14 months of osimertinib treatment, his disease progressed, and next-generation sequencing was performed from a liquid
biopsy of the patient’s blood. An EML4-ALK rearrangement was found and crizotinib was administered. The patient’s lung lesions
continued to progress after one month of crizotinib treatment, and pemetrexed-bevacizumab was initiated. After two cycles of
chemotherapy, the metastatic cancers shrunk, and the patient maintained stable disease at his last follow-up. EML4-ALK
rearrangements can happen in patients with EGFR-positive NSCLC, after acquired resistance to EGFR TKI treatment. The EGFR
T790M and C797G mutations occur in cis is a critical mechanism of resistance to osimertinib therapy. The MAF of EML4-ALK
rearrangements in cancer cells might be a predictive factor for crizotinib treatment.
KEYWORDS Non-small cell lung cancer; EML4-ALK; target therapy; crizotinib

Introduction
The epidermal growth factor receptor (EGFR) is a
transmembrane protein, and upon mutation, is a key driver
gene in non-small cell lung cancer (NSCLC). In Southeast
Asia and China, patients with lung cancer frequently have
high rates of EGFR mutations, especially non-smoking
women1-3. EGFR tyrosine kinase inhibitors (EGFR TKIs),
which reversibly interact with the ATP binding pocket of the
kinase and inhibit cancer proliferation and metastasis, have
shown promising results in patients with EGFR mutation-
positive NSCLC. Patients who are treated with gefitinib or
erlotinib usually acquire resistance, with median progression-
free survival (PFS) durations of 10 to 14 months4,5. About
50%–60% of acquired resistances are induced by the EGFR
T790M mutation, which is a second EGFR point mutation
that involves a threonine to methionine amino acid
substation at position 790 (T790M), is often detected in
tumor cells after disease progression. This mutation enhances
EGFR affinity for ATP, reduces the ability of ATP-
competitive and reversible EGFR TKIs to bind to the EGFR
tyrosine kinase domain, and results in cancer cell resistance
to gefitinib and erlotinib6. The EGFR T790M mutation is the
most common cause of acquired drug resistance for first-
generation EGFR TKIs. Monotherapy with osimertinib has
been associated with a response rate of 61% and a PFS of 9.6
months, with limited skin and gastrointestinal adverse effects,
among patients with EGFR T790M-positive cancers7. The
mechanisms of resistance to osimertinib or other third-
generation EGFR TKIs are very complicated, and the
reported mutation sites and/or rates have substantially varied
among related studies. Phenotypic transformation, new
EGFR point mutation, targets loss, or pathways activation


Correspondence to: Youxin Ji
E-mail: mdji001@gmail.com
Received January 5, 2018; accepted March 20, 2018.
Available at www.cancerbiomed.org
Copyright © 2018 by Cancer Biology &Medicine
Cancer Biol Med 2018. doi: 10.20892/j.issn.2095-3941.2018.0003

(c-Met amplifications, EML4-ALK rearrangement, etc.) have
been the most frequent causes of acquired resistance8,9.
Patients with EML4-ALK rearrangements often manifest
dramatic responses to crizotinib and acquire resistance, with
progression free survivals of about 11 months10-13. ALK gene
point mutations are the primary mechanism of crizotinib
acquired resistance; however, the intrinsic mechanism is
poorly understood14. Here, we report on a patient with
NSCLC with an EML4-ALK rearrangement who experienced
primary resistance to crizotinib treatment.
Case report
The patient was a 66-year-old Chinese man, who was
admitted to our hospital for coughing and blood spiting that
lasted for a month. A chest computer assisted tomography
(CAT) scan showed a 2.5 × 2.5 cm round node on his left
upper lung (Figure 1A). A lobotomy of the left lung and
mediastinal lymph node resections were performed on
November 29, 2012, and a stage IIIA (T1N2M0) lung
adenocarcinoma that metastasized to the left hilar and
mediastinal lymph nodes was histologically confirmed. Four
cycles of adjuvant chemotherapy with cisplatin and
pemetrexed were administered, four weeks after surgery.
Eighteen months later, the patient complained of right upper
abdominal pain, and a positron emission tomography/
computed tomography (PET/CT) scan showed a 3 × 3 cm
mass on the liver and 1.5 × 1.5 cm node on the right adrenal
gland (Figure 1B). A core needle biopsy of the liver lesion
was performed, and a diagnosis of metastatic lung
adenocarcinoma was made by a pathologist. The EGFR gene
panel was examined, in the biopsied tissue, by Amplification
Refractory Mutation System polymerase chain reaction
(ARMS PCR), and an EGFR 19 del was detected. Gefitinib
was administered, and the patient achieved a partial
response. After 18 months of gefitinib treatment, the patient
complained of right leg pain, and his abdominal CAT scan
showed multi-peritoneal lymph nodes metastases. A bone
scan showed right femur metastasis (Figure 1C), and femoral
head replacement to prevent femoral fracture was performed
in December 2015. Bone metastasis was confirmed by the
pathologist, and the EGFR T790M mutation was identified in
the bone tissue. Osimertinib was then administrated, and the
patient achieved a partial response. Fourteen months after
remission, his lung disease progressed (Figure 1D). Next-
generation sequencing (NGS) was performed on the patient’s
blood, and of the 1021 genes that were tested, four genetic
point mutations and an EML4-ALK rearrangement were
found. The mutant allele fractions (MAFs) for EGFR 19 del,
T790M, C797G, and PIK3CA were 2.8%, 4.6%, 2.6%, and
0.9%, respectively, and the EML4-ALK rearrangement rate
was 0.9%. Because the EGFR T790M and C797G mutations
occurred in cis, which would generate resistance to all EGFR
TKIs, crizotinib (Xalkori, Pfizer, USA) was administered.
After one month of crizotinib treatment, the patient’s
condition further deteriorated, and his chest CT scan showed
disease progression (Figure 1E). Chemotherapy with
pemetrexed and bevacizumab was initiated, and the patient
maintained stable disease at his last follow-up on December
20, 2017 (Figure 1F).
A B C
DFE

Figure 1  (A) A CAT scan showed a 2.5 × 2.5 cm, round node on the left upper lung and an irregular mediastinal lymph node. (B) A PET-CT
scan showed liver metastasis. (C) A bone scan showed femoral metastasis, after 18 months of gefitinib treatment. (D) New lesions occurred
after 14 months of osimertinib treatment. (E) Disease progression after crizotinib therapy for one month. (F) Stable disease, after three
cycles of pemetrexed plus bevacizumab therapy.
Cancer Biol Med Vol 15, No 2 May 2018 179

Discussion
Adenocarcinoma and large cell carcinoma account for
50%–60% of lung cancers. About 50% of Eastern Asian and
15% of Caucasian patients with non-squamous NSCLC
harbor EGFR mutations1,15. Furthermore, 4%–7% of patients
with NSCLC have EML4-ALK rearrangements, and ROS1
rearrangements and BRAF V600 mutations occur at lower
frequencies16,17.
Previous studies have established EGFR TKIs as the
mainstay treatment for patients with NSCLC harboring
EGFR activating mutations18. However, there is an eventual
loss of TKI efficacy due to development of acquired
resistance. The EGFR T790M mutation is the leading cause
for resistance to first generation EGFR TKIs, but for third
generation EGFR TKIs, the resistance mechanism is much
more complicated. Hence, the discovery of new drugs that
can overcome resistance to third generation EGFR TKIs is of
critical importance for prolonging the survival of patients
with NSCLC19,20.
The EML4-ALK translocation involves a fusion of the N -
terminus of EML-4 with the kinase domain of ALK, to
produce the EML4-ALK fusion gene. A series of research has
shown that this translocation involves several variations, all
of which lead to catalytically active kinase fusion protein
variants, and experiments in animal models have proven that
the kinase activity is required for carcinogenicity21,22. ALK
rearrangement-positive advanced NSCLC has had high
response rates to crizotinib treatment; however, the exact
mutation allele fraction (MAF) threshold that is needed to
achieve a treatment response has not been reported in
English23. In this case, the patient’s lung lesions progressed
quickly after the initiation of crizotinib treatment. This might
be because the patient harbored an EGFR 19 del, T790M
mutation, and C797G mutation, as primary tumor driver
genes, or because the EML4-ALK rearrangement MAF was
too low.
For patients harboring EGFR T790M and C797G
mutations, we must distinguish whether the two mutations
occurred in cis or in trans. In this patient, three EGFR point
mutations occurred in cis, which indicated that the patient
would be resistant to all EGFR TKIs24,25. Previous data has
shown that checkpoints inhibitors had limited effects on
EGFR-mutated lung cancers, because the tumor cells either
lacked PD-L1 or expressed PD-L1 at low levels26-28.
Chemotherapy was the optimal choice, and low toxicity
drugs with pemetrexed were used, since the patient had a
long treatment history, and his performance status (PS),
according to the Eastern Cooperative Oncology Group
(ECOG) scoring system, was 2.
The formation of new blood vessels, known as
angiogenesis, is a fundamental event in the process of tumor
growth and metastatic dissemination. The vascular
endothelial growth factor (VEGF) and its receptors play an
essential role in tumor proliferation29-31. Bevacizumab, a
recombinant humanized monoclonal antibody, combined
with VEGFA, attenuates VEGFA-dependent tumor blood
vessels formation, normalizes tumor blood vessels, prompts
tumor cell apoptosis, and shrinks tumors. This patient
presented multi-organ metastasis, in which the VEGF
signaling pathway might play an important role.
Conclusions
The EGFR T790M and C797G mutations occurred in cis
were the critical mechanisms of resistance to osimertinib.
The EML4-ALK rearrangement was found in this patient
with EGFR-positive NSCLC after acquired resistance to
EGFR TKI treatment. The EML4-ALK MAF in cancer cells
might be a predictive factor for crizotinib treatment.
Chemotherapy was the optimal treatment method for this
patient, who harbored EGFR T790M and C797G mutations
in cis and had a synchronous EML4-ALK rearrangement.
Conflict of interest statement
No potential conflicts of interest are disclosed.
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Cite this article as: Zhang L, Li Y, Zhang S, Gao C, Nie K, Ji Y, et al. Primary
resistance to crizotinib treatment in a non-small cell lung cancer patient with
an EML4-ALK rearrangement: a case report. Cancer Biol Med. 2018; 15: 178-
81. doi: 10.20892/j.issn.2095-3941.2018.0003
Cancer Biol Med Vol 15, No 2 May 2018 181