Perspectives
Snapshot
Yale Cancer Center Precision Medicine Tumor Board:
new technology, new drugs, and the value of repeat testing
A 41-year-old woman who had never smoked was found
to have a 2·1 cm nodule in the left lower lobe of the lung
without nodal or distant metastases in February, 2009.
She underwent a left lower lobectomy, and pathological
evaluation showed a 2·5 cm moderately differentiated
adenocarcinoma with immunohistochemistry positive
for TTF-1 and CK7, and negative for CDK2, CK20, and
mammoglobin, consistent with a non-small cell lung cancer
(NSCLC). Invasion of the visceral pleura was identified
and one hilar lymph node was positive for metastasis.
Assessments of EGFR exons 18–21 by Sanger sequencing
and KRAS codons 12 and 13 by primer extension assays
showed no mutations. The patient received four cycles
of adjuvant chemotherapy. In June, 2012, surveillance
imaging showed an enlarged right lung nodule that was
resected to reveal a 0·5 cm adenocarcinoma. In 2014,
the patient developed additional right-sided nodules
and received stereotactic body radiotherapy. Imaging in
June, 2015, showed multiple bilateral lung nodules, which
resulted in a recommendation to initiate systemic therapy.
Multiple methods were used to test the 2012 resected
tumour specimen for molecular alterations. Targeted
PCR-based assays did not detect any common mutations
in seven genes. All exons of 409 cancer-related genes
were then examined by next-generation sequencing,
using a commercial assay (Comprehensive Cancer Panel,
ThermoFisher Scientific, Waltham, MA, USA), in which
highly multiplexed PCR are employed for target selection,
followed by amplicon sequencing on an Ion Torrent
platform (ThermoFisher Scientific, Waltham, MA, USA); only
subclonal mutations in TGFBR2 and CTNNB1 were detected.
Fluorescence in situ hybridization (FISH) was used to test
for genomic rearrangements involving ALK, ROS1, and RET.
No ALK rearrangement was detected. FISH did not provide
evidence for a ROS1 rearrangement, although interpretation
was complicated by chromosome 6 monosomy. RET status
was not evaluable due to technical failure of the assay.
Since no driver alteration was detected, the patient was
treated with pembrolizumab as part of a clinical trial and
had prolonged disease stability. In April, 2017, she had
disease progression in the lungs, prompting transition to
a trial with combination immunotherapy. In April, 2018,
her disease progressed to the liver and a left supraclavicular
lymph node. The left supraclavicular lymph node was
biopsied for further analysis. Immunohistochemical
staining revealed PD-L1 positivity in both the tumour cells
(1–50%) and immune cells (>1%).
www.thelancet.com/oncology Vol 21 March 2020
Molecular pathology results
The supraclavicular lymph node biopsy, containing an
estimated 20% malignant cells, was analysed in the
Yale Tumor Profiling Laboratory using the Oncomine
Comprehensive Assay (version 3; ThermoFisher
Scientific). This amplicon-based next-generation
sequencing assay examines tumour DNA for mutations
in 135 genes and amplifications of 43 genes, and also
examines tumour RNA for the presence of gene fusion
transcripts involving 44 oncogenic driver genes. An
NCOA4-RET gene fusion was found at a low level; no
other alterations were detected. Rearrangement of the
RET genomic locus was subsequently confirmed by FISH.
Discussion
RET (ret proto-oncogene) encodes a receptor tyrosine
kinase that functions in multiple tissues during normal
renal, neuroendocrine, and nervous system development.
Oncogenic activation of the RET tyrosine kinase can
occur via mutation of the coding sequence (often found
in medullary thyroid carcinoma), or by chromosomal
rearrangement that fuses the kinase domain of RET to a
variety of partner genes (often found in papillary thyroid
cancer). RET fusions are also found in 1–2% of NSCLCs,
primarily adenocarcinomas in patients with little or no
smoking history. To date, RET fusions involving at least
15 different partner genes have been described in NSCLC.
NCOA4 is the third most common RET fusion partner
in NSCLC, after KIF5B and CCCD6. Generally, RET fusion
Department of Medicine,
Medical Oncology (NH, JPE,
SNG, SBG), and Department of
Pathology (ZW, JLS, KEF),
Yale School of Medicine,
New Haven, CT 06510, USA
sarah.goldberg@yale.edu
NH and ZW contributed equally
All authors interpreted the data,
reviewed and gave approval for
the final manuscript. NH, ZW,
and SBG wrote the manuscript
SBG reports grants and personal
fees from AstraZeneca,
and personal fees from
Boehringer Ingelheim,
Bristol-Myers Squibb, Eli Lilly
Genentech, Spectrum, and
Amgen, outside the submitted
work. NH, ZW, JPE, JLS, SNG,
and KEF declare no competing
interests
For more on RET fusions in
NSCLC see J Clin Oncol 2012;
30: 4352–59
For more on RET fusion partners
in NSCLC see Front Physiol 2019;
10: 216 and J Clin Oncol 2012;
35: 1403–10
For more on molecularly driven
therapies see JAMA Oncol 2018;
4: 1093–98
Kateryna Kons/Science Photo Library
History
343
Perspectives
For more on NSCLC oncogenic
alterations see Ann Oncol 2019;
30 (suppl 5): v602–60,
Lancet Oncol 2016; 17: 1653–60,
Expert Opin Investig Drugs 2018;
27: 363–75, and Lancet Oncol
2017; 18: 1307–16
For more on RET TKIs for lung
cancer see J Thorac Oncol 2019;
14: S6–7 and J Clin Oncol 2018;
37 (suppl 15): 9008
344
proteins are oncogenic because the domain contributed by
the partner gene mediates dimerisation, mimicking ligandinduced dimerisation of the native receptor, and leads to a
constitutively active RET kinase domain within the fusion
protein. This activated RET kinase domain continually
initiates signal transduction cascades, such as those of the
RAS and PI3K/AKT pathways, resulting in sustained proproliferative intracellular signalling.
Since this patient was young with no history of
smoking, the likelihood that her tumour harboured
an actionable molecular driver was increased, which
prompted physicians to pursue repeated tumour profiling
over time. For advanced NSCLC, the spectrum of relevant
genetic alterations and of available testing methods has
changed in recent years, and will probably continue to
evolve in parallel with advances in knowledge of genomic
underpinnings of cancer, drug development, and tumour
profiling assay technologies. The full promise of precision
medicine remains to be realised, since molecular profiling
does not identify therapeutic options in most patients with
advanced cancer. However, the number of patients with
advanced cancer who are eligible for a US Food and Drug
Administration (FDA)-approved molecularly driven therapy
has steadily increased, from 10·5% in 2006, to 15·4% in
2018, indicating the importance of continuously updating
testing platforms.
The growing number of known molecular drivers in
NSCLC with associated targeted therapies raises the
question of when a molecular pathology report showing
no actionable genetic alterations should be accepted
as final and satisfactory. In the early days of targeted
therapy for NSCLC, EGFR mutational analysis was
sufficient. Assessment for ALK and ROS1 gene fusions
became necessary with the development of effective
therapies for those alterations. However, at present,
BRAF mutations in lung cancer are targetable with an
FDA-approved therapy (dabrafenib and trametinib), and
promising results from recent clinical trials suggest that
oncogenic alterations in NTRK1/2/3, MET, and RET are
all therapeutically actionable. Our patient’s experience
reflects the expanding possibilities for NSCLC therapy.
Although RET fusions have been described in papillary
thyroid cancer since 1990, RET fusions in NSCLC were
first reported in 2012. Soon thereafter, treatment options
emerged. This patient’s cancer was indolent, enabling
the opportunity for repeated testing over several years.
By 2018, RET fusion assessment had become particularly
relevant considering emerging data on the success of
novel RET tyrosine kinase inhibitors.
Various methods can detect oncogenic gene fusions in
solid tumours, including FISH, immunohistochemistry,
RT-PCR, and next-generation sequencing, each with
strengths and limitations. At present, many academic and
commercial laboratories offer targeted next-generation
sequencing panels that assess hundreds of cancer-related
genes for mutations and gene amplifications and that
detect dozens of common gene fusions, including those
known to occur in lung cancer. Some next-generation
sequencing assays identify gene fusions by sequencing
intronic DNA regions that contain the breakpoints of
recurrent chromosomal rearrangements. Others assays,
such as the Oncomine assay, identify gene fusions in the
tumour RNA by sequencing the products of a multiplex RTPCR reaction designed to amplify specific fusion junctions
among transcripts generated by a large set of possible gene
fusions. Because oncogenic gene fusions are expected to be
highly expressed, RNA-based methods might have greater
sensitivity to detect gene fusion events compared with
DNA-based methods in samples with low malignant cell
content.
In this case, the initial molecular testing examined only
EGFR and KRAS, and the next-generation sequencing
panel used in 2012, did not assess gene fusions.
Additionally, although FISH testing showed no evidence
of ALK or ROS1 rearrangement, the RET FISH assay at that
time did not yield interpretable results due to technical
failure. In 2018, a RET gene fusion was detected by
next-generation sequencing at a low level in a sample
containing a low proportion of malignant cells. The nextgeneration sequencing result was then confirmed by a
RET FISH assay.
Precision Medicine Tumor Board recommendation
Clinical trial investigating a RET inhibitor.
Clinical follow-up
The patient was started on a clinical trial with a novel
RET tyrosine kinase inhibitor and had substantial disease
shrinkage that continues 1 year after initiating therapy.
Conclusion
Considering recent progress in the treatment of lung
cancer, it is crucial to use assay methods that provide
a comprehensive analysis for all relevant molecular
alterations. In cases such as this, in which previous testing
did not provide a definitive assessment of all actionable
alterations, repeat testing is indicated and can lead to
effective therapies for patients.
Navid Hafez, Zenta Walther, Joseph P Eder, Jeffrey L Sklar,
Scott N Gettinger, Karin E Finberg, *Sarah B Goldberg
www.thelancet.com/oncology Vol 21 March 2020