23 Lung cancer Flashcards
What does SCLC stand for in lung cancer?
1) Small cell lung cancer;
2) About 15% of all lung cancer;
3) It is sometimes called oat cell cancer. This type of lung cancer tends to grow and spread faster than NSCLC. In most people with SCLC, the cancer has already spread beyond the lungs at the time it is diagnosed.
What does NSCLC stand for in lung cancer?
1) non-small cell lung cancer;
2) 85% of all lung cancer;
3) NSCLC is any type of epithelial lung cancer other than small cell lung cancer (SCLC);
4) The most common types of NSCLC are:
– squamous cell carcinoma (SQCC, 25%);
– large cell carcinoma (10%);
– adenocarcinoma (ADC, 40%);
– adenosquamous carcinoma;
– other less common subtypes
what are the most common types of NSCLC?
1) squamous cell carcinoma (SQCC, 25%);
2) large cell carcinoma (10%);
3) adenocarcinoma (ADC, 40%);
4) adenosquamous carcinoma;
5) other less common subtypes
What three markers can reduce the number of NSCLC-not otherwise specified (NOS) diagnoses to less than 5-10%?
1) thyroid transcription factor-1 (TIF-1);
2) a mucin stain;
3) either p63 or p40
what is Lepidic adenocarcinoma?
Lepidic adenocarcinoma represents a histologic pattern of NSCLC that characteristically arises in the lung periphery with tracking alongside pre-existing alveolar walls.
what is Acinar adenocarcinoma?
1) Acinar adenocarcinoma is a histological subtype of gland-forming cancer.
2) Acinar adenocarcinoma is diagnosed when cuboidal and/or columnar shaped malignant cells in the neoplastic tissue form acini and tubules.
3) Acinar adenocarcinoma is a common form of cancer occurring in the lung and prostate gland.
Papillary adenocarcinoma of lung
1) Papillary adenocarcinoma of lung is a variant where the major component shows papillary growth pattern, often with complex secondary and tertiary branching structure.
2) The tumor cells are arranged in multiple layers around true fibrovascular cores.
Micropapillary-predominant adenocarcinoma (MPA)
1) Micropapillary-predominant adenocarcinoma (MPA) is evaluated as a high-grade subtype with a poor prognosis.
2) The micropapillary pattern in lung adenocarcinoma is characterized by small papillary tufts lying in alveolar spaces or in spaces encased by connective tissues, with the tufts having no fibrovascular core,
What biomarkers are used as the basis for personalized medicine in lung cancer patients?
EGFR (epidermal growth factor receptor);
ALK (anaplastic lymphoma kinase)
What is EGFR?
1) epidermal growth factor receptor;
2) aka. ErbB-1;
3) EGFR is a transmembrane receptor tyrosine kinase (RTK)
4) a 170-kDa membrane glycoprotein
Where is EGFR commonly overexpressed?
EGFR is commonly overexpressed in a wide range of cancers from:
1) Bladder
2) Brain
3) Breast
4) Colorectal
5) Head and neck
6) Lung
7) Pancreas
EGFR dimer partners
EGFR forms homodimers and heterodimers with other members of the ErbB family receptors, including:
1) HER2/c-neu (ErbB-2)
2) HER3 (ErbB-3)
3) HER4 (ErbB-4)
epiregulin, aka EREG
1) Epiregulin (EREG) belongs to the epidermal growth factor (EGF) family;
2) binds to the epidermal growth factor receptor (EGFR);
3) regulates the immune response of the host during infections.
Heparin-binding EGF-like growth factor (HB-EGF)
Heparin-binding EGF-like growth factor has a role in Angiopoietin-mediated recruitment of vascular smooth muscle cells
Amphiregulin is a key molecule among epidermal growth factor receptor (EGFR) ligands and may play a pivotal role in the development or maturation of placenta
EGFR ligands
examples of EGFR ligands:
1) EGF;
2) TGFα (transforming growth factor alpha);
3) amphiregulin (amp-hire-gulin);
4) betacellulin (beta-cell-ulin);
5) heparin-binding like EGF factor (HB-EGF);
6) epiregulin, aka EREG
How is EGFR activated?
1) The binding of ligand to EGFR generates a shift in conformation, enabling EGFR to dimerize, which in turn activates the cytoplasmic intracellular tyrosine kinase (TK) domain to become activated through autophosphorylation in the C-terminal domain of EGFR.
2) Ligand binding –> EGFR dimerization–> TK autophosphorylation –> activation
EGFR downstream signaling pathways
1) RAS/RAF/MAPK;
2) PI3K/AKT;
3) JNK;
4) JAK/STAT
Function of EGFR activation
leads to cell proliferation and survival
Function of EGFR mutations in cancer development
1) Extracellular EGFR mutations lead to constitutive activation of the receptor independent of the ligand binding;
2) EGFR mutations can lead to EGFR overexpression;
3) overactivated EGFR have been associated with carcinogenesis;
4) ATP-binding cleft mutations may confer sensitivity to targeted EGFR-TKIs.
Domains of EGFR
1) an extracellular ligand-binding domain;
2) a transmembrane segment;
3) a large intracellular domain including a protein tyrosine kinase (TK) domain
Which exons encode TK domain of EGFR?
exon 18-21
Classification of EGFR TK domain mutations
1) in-frame deletions of exon 19;
2) missense (point) mutations in exons 18-21;
3) insertion in exon 20
About 90% of mutations associated with EGFR-TKI sensitivity located in which exons of EGFR?
1) Exons 19 and 21;
2) Exon19 mutations:
– LREA deletions (45%): leucine, arginine, glutamic acid, alanine;
– VAIKEL insertions (1%): valine, alanine, isoleucine, lysine, glutamic acid, leucine
3) Exon 21 mutations:
– L858R (40%);
– L861X (2%)
The most common acquired EGFR mutation following TKIs treatment is in lung cancer?
1) Exon 20 EGFR T790M (threonine to methionine);
2) EGFR T790M is a missense mutation;
3) 50% of patients develop TKIs resistance to first generation TKIs therapy showed EGFR T790M, and these patients have preexisting sensitizing EGFR mutations.
Which ethnic group is associated with EGFR mutations?
female, East Asian
TTF1
1) Thyroid transcription factor 1;
2) TTF1 is a highly specific marker for primary lung adenocarcinomas
EGFR mutations are more commonly associated with what clinical characteristics?
1) Female East Asian (30-50% prevalence); Caucasian 10-20%;
2) nonsmokers or light smokers of <10 pack-years;
3) TTF1 IHC positive of adenocarcinoma;
4) nonmucinous morphology
EGFR mutations present more in which type of lung cancer?
NSCLC, adenosquamous carcinomas;
rare in SCC (squamous cell carcinoma)
Effect of preexisting EGFR mutations on TKIs therapy
EGFR T790M results in some hindrance of the kinase receptor domain and cause difficulty for TKI binding while maintaining the ability to bind ATP.
This reduces the TKIs potency.
What novel agents are designed for patients with EGFR T790M but have a TKIs sensitizing mutation?
AZD9291, HM61713, CO-1686
The % of patients with TKIs resistance has focal amplification of c-MET?
1) c-MET amplifications in 5-10% of cases;
2) leading to reactivation of the signaling pathway for cell proliferation despite of the presence of EGFR inhibition;
3) Thus, multiple kinase inhibitions should be considered for a more durable treatment.
EGFR overexpression effect in lung cancer treatment
1) Overexpression of EGFR did not show increased TKIs response, thus overexpression of EGFR testing is not recommended for selection of targeted therapy.
2) EGFR IHC is not recommended neither
EGFR overexpression detecting method
1) FISH;
2) silver in situ hybridization;
3) real-time qPCR
what is ALK?
1) Anaplastic lymphoma kinase;
2) aka CD246;
3) a transmembrane TK receptor;
4) plays a role in the development of the peripheral nervous system;
5) ALK gene is located at chr. 2p23
ALK was first identified as a fusion partner of which protein?
ALK was first identified as a fusion partner of fusion partner with nucleophosmin (NPM1) in a subset of anaplastic large-cell lymphoma.
ALK downstream signaling pathways?
1) RAS/RAF/MAPK;
2) PI3K/AKT;
3) JAK-STAT
% of NSCLC with ALK fusion with other genes?
5-7%
The most common gene fusion with ALK in lung cancer?
1) EML4-ALK;
2) inversion of EML4
Clinical features associated with ALK rearrangements?
1) nonsmoking or light-smoking history;
2) younger age;
3) histologic characteristics: ADC histology, mucinous morphology, signet-ring cell morphology
ALK rearrangements are mutually exclusive of other oncogenic driver mutations.
Cancer with ALK rearrangement show dependency on the continued signaling of the fusion protein and are thus highly susceptible to targeted therapy.
What are the other ALK fusion partners in lung cancer?
KIF5B;
TFG;
KLC1;
In all cases, the genomic breakpoint of ALK is well conserved, resulting in the fusion of the partner protein to the intracellular domain of ALK.
The effect of EML4-ALK fusion in lung cancer?
uncontrolled cell proliferation.
What is the first generation of ALK inhibitor?
crizotinib;
Crizotinib resistance developed within 1-2 years in ALK fusion-positive NSCLC.
Most common ALK mutations in lung cancer
1) ALK TK domain L1196M;
2) This mutation causes steric hindrance at the binding site, resulting in decreased potency of response to crizotinib.
Other ALK mutations of the TK domain in lung cancer that lead to TKIs resistance
ALK G1269A, G1202R, S1206Y, F1174C/L, D1203N
ALK domain away from the binding site in lung cancer that lead to TKIs resistance
ALK threonine insertion at 1151;
ALK C1156Y, L1152R
ALK inhibition in lung cancer lead to what other kinases activation?
1) EGFR or cKIT amplification–>activate cell signaling pathways for cell proliferation
2) Thus need to inhibit the secondary kinase activation
What is KRAS?
1) Kirsten rat sarcoma viral oncogene homolog;
2) KRAS is an intracellular GTPase that is tethered to the cell membrane
Signaling pathways activated by KRAS?
1) PI3K/AKT;
2) RAF/MEK/ERK;
3) RLF/RAL;
4) leading to cell proliferation and cell survival when KRAS is activated.
How the KRAS activity is stopped?
KRAS has intrinsic enzymatic activity to cleave GTP to GDP, stopping the downstream signaling pathways.
Effect of KRAS mutations at codons 12 and 13 of exon 2.
Mutations in codons 12 and 13 of exon 2 of KRAS, resulting in an inability to hydrolyze GTP to GDP, leaving KRAS constitutively activated.
% of lung cancer ADC with KRAS mutations
25-35%
Is KRAS downstream of EGFR?
Yes
KRAS and EGFR mutations are mutually exclusive in lung cancer.
KRAS and EGFR mutations are mutually exclusive in lung cancer.
Clinical feature of KRAS mutations in lung cancer
1) associated with smoker;
2) mucinous morphology;
3) 5-yrs survival is not different in KRAS mutated patients vs. wild type KRAS in NSCLC
Mechanisms of resistance to crizotinib therapy in lung cancer
1) Acquired resistance mutations in ALK such as
L1195M, G1269A;
2) ALK fusion amplified copy number gains (CNG)
– 47% of cases;
– activating an alternate pathway (EGFR, CKIT,
MET);
3) evolution of a ALK-TKI resistance mutation.
ROS1 gene
1) ROS1 gene is located on the long arm of chr. 6 (6q22);
2) ROS1 gene encodes a type-1 integral membrane protein with TK activity;
3) Protein is ROS proto-oncogene 1;
4) ROS1 is a member of the insulin receptor family
Function of ROS1
Activation of ROS1 results in growth and differentiation of a cell through the MAPK pathway and phosphorylation of RAS.
Role of ROS1 in lung ADC?
1) ROS1 rearrangements have been identified in
~2.5% lung ADC;
2) ROS1 most common fusion partner is CD74
– t(5;6)(q32:q22)
Clinical features of ROS1 rearrangement NSCLC patients
1) Younger age with nonsmoking or light smoking history;
2) Asian;
3) ADC history
How to detect ROS1 rearrangements in lung cancer
FISH with dual break-apart probes
ROS1-rearranged and ALK-rearranged cancers are mutually exclusive.
BRAF gene
1) BRAF gene is located on the long arm of chr. 7
(7q34);
2) BRAF gene encodes for the serine/threonine
protein kinase, B-raf;
3) B-Raf is a member of the Raf kinase family, and is downstream target of Ras;
4) Role in MAPK/ERK signaling pathway
What is the common BRAF mutations in NSCLC?
BRAF V600E;
it is also common in melanoma
Clinical features of BRAF V600E in lung cancer?
Younger, nonsmoking female;
Show micropapillary ADC pattern
Clinical features of BRAF non-V600E in ling cancer?
smoking history;
Caucasian
Multiple BRAF inhibitors are available with specific targeted affinity for the BRAF V600E NSCLC.
Patients with non-V600E BRAF mutations that showed resistance to BRAF inhibition, what they should do next?
try MEK inhibitors, such as Selumetinib-AZD6244
Disadvantages for Sanger sequencing for tumor gene mutations testing?
1) The requirement for 40-50% tumor cellularity in the sample, leading to higher false negative.
2) FFPE can have artifactual mutations due to artifacts of deamination at cytosine and adenine (with transition to uracil and hypoxanthine); to avoid this, treat the 1 ug DNA with uracil-N-glycosylase
Advantages in using NGS for mutation detection.
1) NGS showed improved sensitivity of mutation detection in cases with tumor cellularity less than 40%;
2) High throughput;
3) Detect multiple forms of mutations: SNP, copy number alterations, rearrangements, InDel
List of ultrasensitive testing platforms that offer the advantages of high caseload throughput and rapid turnaround time
1) Amplification refractory mutation system;
2) length analysis;
3) restriction length polymorphism;
4) real-time PCR;
5) high-resolution melting curve analysis;
6) single-base extension genotyping;
7) mass spectrometry;
8) denaturing high-performance liquid
chromatography;
Advantage: the ability to detect very low caner cellularity (<10%)
Disadvantage: detection of only a limited scope of very specific mutations.
What kinds of lung cancer patient should be tested for all individual EGFR mutations (1% allele frequency), according to CAP/IASLC/AMP?
1) ADC histology;
2) for which the presence of an ADC component can not be excluded
EGFR mutation-specific IHC can be used for which EGFR mutations?
1) EGFR L858R on exon 21:
–specificity: 97.8%;
–sensitivity: 75.6%;
2) EGFR the exon 19 15bp detection:
–specificity: 95.5%;
–sensitivity: 42.2%;
3) Low sensitivity but allow rapid identification of a TKI-sensitizing mutation to facilitate early initiation of treatment
CAP/IASLC/AMP recommends what gene mutations should be tested in lung cancer with ADC or ADC component?
EGFR mutations;
ALK rearrangements
How is ALK rearrangements detected in lung cancer?
1) by a dual-labeled break-apart FISH assay;
2) FDA approved, Abbott Molecular;
3) Detection of a fusion product is identified when the SpectrumOrange-labeled 5’ telomeric end of ALK is identified separate from the SpectrumGreen labeled 3’ centromeric end;
4) Cases are considered positive if 15% or more than 50 tumor nuclei are identified with the split signal;
4) Formalin-fixed specimens are preferred since alcohol fixatives may interfere with FISH probes.
When a diagnosis cannot be established based on morphologic features alone, what is the next step to take for lung cancer diagnosis?
1) To establish ADC lineage: TTF-1 or mucin stain;
2) To establish SCC lineage: p63/p40 or CK5/6 stain
When to use molecular testing in lung cancer diagnosis?
1) For patients with advanced-stage disease (stage IV) who are suitable for targeted therapy;
2) or at the time of recurrence;
3) or progression in patients who presented at an earlier stage.–> archived specimen can be used
Some studies have shown that mutations can be detected with how many tumor cells?
1) 100-400 tumor cells;
2) 1-3% tumor DNA content
What is the criteria for ideal sample for molecular testing?
good preservation of tumor DNA, high tumor cellularity, limited necrosis and mucin
What fixatives might interfere with molecular testing?
1) fixatives containing heavy metals: Zenker, B5, B-plus, acid-zinc formalin;
2) acidic decalcifying solutions may result in extensive DNA fragmentation
What is CTC?
1) circulating tumor cell;
2) used to monitor the presence and genetic evolution of solid tumors;
3) CTCs can be detected in up to 70% of patients with metastatic disease;
4) CTCs may play a role in real time monitoring of treatment response and evolution of acquired resistance to TKIs;
5) currently, the presence of specific mutations in NSCLC CTCs have good specificity but poor sensitivity.
What is ctDNA?
1) circulating tumor DNA;
2) DNA fragments of 140-170bp;
3) ctDNA can be found in plasma or cell-free fraction of blood.
How much amplifiable DNA present in 1 mL of blood?
between 30 and 40 µg/mL
of which 0.01-0.1% is ctDNA
ctDNA can be identified in the plasma of patients who have only localized disease and do not show CTCs.
Unlike CTCs, NGS analysis of ctDNA from NSCLC represents both a specific and highly sensitive biomarker that can be used to detect cancer burden, oncogenic mutations, and TKI-resistance mutation subclones before clinically or radiologically apparent.
EGFR TKIs
1) first generation 4-anilinoquinazoline small molecule reversible TKIs:
— gefitinib;
— erlotinib;
— about 60-80% of EGFR mutation-positive chemotherapy-naive patients show response;
— all patients eventually progress due to acquired drug resistance.
2) 2nd-generation quinazoline-based small-molecule irreversible TKIs:
— afatinib:
— increased PFS for patients with 1st-generation TKI resistance;
— Afatinib used in combination with paclitaxel improves the response in patients with 1st-generation TKIs resistance;
— dacomitinib;
— bind both wild type and mutated EGFR;
3) 3rd-generation non-quinazoline-based TKIs:
— AZD9291;
— rociletinib;
— HM61713;
— preferential activity against EGFR T790M than the WT;
4) monoclonal antibodies target EGFR by binding to the extracellular domain of EGFR:
— cetuximab;
— failed in clinical trial to be used by itself
How can Afatinib improves the response in patients with 1st-generation TKIs resistance to EGFR-targeted therapy in lung cancer
1) Afatinib used in combination with paclitaxel;
2) Afatinib used with centuximab:
— afatinib targets the phosphorylated EGFR;
— centuximab affects total EGFR protein expression
ALK-targeted therapy in lung cancer
1) 1st-generation ALK-TKIs:
– crizotinib: also inhibits ROS1;
– 50-60% of patients with ALK-positive NSCLC with PFS of 8-9 months;
– all patients develop acquired resistance to crizotinib within 1 year.
2) 2nd-generation ALK-TKIs:
– ceritinib: 20-fold potency than crizotinib, response rate 55-56% in crizotinib-resistant patients;
– alectinib: activity against ALK L1196M and G1269A resistance mutations, response rate 93.5% in crizotinib-naive patients;
– AP26113: dual TKI with activity against native ALK, L1196M ALK, mutated EGFR, and T790M EGFR;
– greater selectivity or potency for the ALK-TK domain to address the issue of acquired resistance
KRAS/MEK-targeted therapy
1) No specific targeted agents are available for KRAS-mutated NSCLC
2) New agents targeting downstream MEK1/MEK2 have shown promise in phase 2 trials.
– MEK inhibitor: selumetinib
