Lung Cancer: Assessment and Treatment

Introduction

Lung cancer is the third most common cancer and the leading cause of cancer-related death in the United States. Two Main Types:

• Small cell lung cancer: 5-year overall survival (OS) = 7%
• Non-small cell lung cancer (NSCLC): 5-year OS = 25%
• Includes: adenocarcinoma, squamous cell carcinoma, large cell carcinoma

Initial Diagnosis

Staging

Staging provides information regarding tumor size and extent the tumor has spread throughout the body.

Clinical Workup

• Imaging: X-ray, MRI, CT and/or PET scan
• Including the brain, bones and liver
• Lung Biopsy: If diagnostic biopsy is an option, enough tissue should be collected to allow for biomarker assessment.

Tumor Molecular Profiling

Tumor Molecular Profiling tissue and/or blood biomarkers help guide and optimize a patient’s treatment decisions and outcomes. It is important for tumor biomarker testing to be performed at diagnosis and repeated at time of progression to assess potential mechanisms of acquired resistance, including additional driver mutations that may have developed due to therapy.

• FISH (fluorescent in situ hybridization): Rapid technique that provides information about tumor DNA.
  • Limitations include tissue depletion, being more difficult to interpret clinically, and not providing information related to fusion partners or positions.
• IHC (immunohistochemistry): Rapid low-cost technique that provides information related to transcribed and translated events (protein information).
  • Limitations include tissue depletion, being less well-validated, and not providing information related to fusion partners or position.
• RT-PCR (reverse-transcriptase polymerase chain reaction): Rapid technique with high sensitivity and specificity that provides information related to RNA and can detect specific fusion partners.
  • Limitations include specific primer sets being required for each fusion, inability to detect novel fusion partners, difficulty in extracting high-quality RNA from tumor, and limited multiplexing ability.
• NGS (next generation sequencing — including RNA and DNA): Technique allows for multiplexing to maximize information obtained from single tissue sample. NGS provides information about targeted DNA and/or RNA locations and/or the whole genome in a single test. It can provide information about fusion partners and identify position.
  • Limitations include higher cost and longer wait time for results.
• Liquid Biopsy: Noninvasive method for assessing circulating tumor-derived materials — cell-free DNA — from plasma in patients at advanced stages where biopsy is not an option.
  • Limitations include lack of reliability in clinical setting.

Current Oncogenic Biomarkers in Lung Cancer

The next sections will delineate current oncogenic biomarkers in lung cancer, reviewing the background, clinical features, detection methods, most common alterations and FDA-approved therapies.

EGFR: Epidermal Growth Factor

Background

Transmembrane receptor tyrosine kinase protein involved in activation of various signaling pathways, including PI3K/Akt. Binding of EGFR to its ligand results in dimerization and tyrosine autophosphorylation leading to cell proliferation and maintenance. Alterations resulting in overexpression of EGFR are associated with tumorigenesis, reduced survival, lymph node metastasis and poor chemosensitivity. EGFR mutations occur predominately in exons 18-21 and are identified in approximately 10-15% of patients with NSCLC.

Clinical Features Associated

Females, minimal/no smoking history, adenocarcinoma histology, Asian ethnicity

Detection Methods

NGS, FISH, PCR, liquid biopsy (when tissue biopsy unavailable)

Most Common Alterations

• Sensitizing (Classical) Mutations: These mutations account for approximately 90% of EGFR mutations in NSCLC. Sensitizing mutations are associated with high sensitivity to and prolonged progression-free survival (PFS) with targeted EGFR tyrosine kinase inhibitors (TKIs) compared with chemotherapy. The most common mutations include:
  • Exon 19 Deletions destabilize the inactive form of EGFR, causing it to remain in the active conformation. These deletions account for around 45% of all EGFR mutations. The most common frame shift deletion is delE756-A750, followed by delL747_P753insS and delL747_A750insP.
  • Exon 21 Point Mutations lock EGFR in a constitutively active conformation and account for 40% of EGFR mutations. The most common exon 21 mutation is L858R.
• Uncommon Mutations: Rare EGFR mutations account for roughly 5-10% of EGFR mutations. The location of these mutations can have significant effects on sensitivity to EGFR TKIs. The most common include exon 20 S7681 (3% EGFR mutations), exon 21 L861Q (1%) and exon 18 G719X (1%).
• Acquired Resistance Mutations: These mutations occur after treatment with TKI therapy. The most common post-first- and second-line EGFR TKI acquired resistance mutation is exon 20 T790M. The most common post-third generation EGFR TKI therapy mutation is exon 20 C797S, which accounts for 10-26% of resistance mutations following osimertinib (Tagrisso, AstraZeneca) therapy.

FDA-Approved Targeted Therapies:

NCCN guidelines recommend osimertinib as first-line therapy for EGFR-sensitizing mutations. Additionally, targeted therapy is recommended prior to chemotherapy or immunotherapy.\

Erlotinib (Tarceva, Roche):

• First-generation EGFR TKI granted accelerated approval by the FDA in 2004 for treatment of patients with locally advanced or metastatic NSCLC who progressed on at least one prior chemotherapy regimen. Approval was based on a single randomized trial of patients with stage IIIB or IV NSCLC who failed one or two prior chemotherapy regimens. A response rate of 8.9% and median OS of 6.7 months was found compared with 4.7 months in the placebo arm. This approval was later expanded by the FDA to include maintenance therapy for patients with locally advanced or metastatic NSCLC who had not progressed after first-line chemotherapy. In 2013, FDA expanded approval of erlotinib further into the first-line setting to include patients with metastatic NSCLC with eitherEGFR exon 19 deletions or exon 21 L858R substitution. This approval was based on a phase 3 clinical trial (EURTAC) with PFS of 10.4 months compared with 5.2 months with chemotherapy.
• Most Common Adverse Events: diarrhea, rash, nausea, vomiting, cough, shortness of breath, decreased appetite

Gefitinib (Iressa, AstraZeneca):

• First-generation EGFR TKI granted accelerated approval by the FDA in 2003 for patients with advanced stage NSCLC who progressed on both platinum-based and docetaxel chemotherapies prior to biomarker screening for EGFR. The approval was based on two phase 2 clinical trials— IDEAL 1 and 2 — with an objective response rate (ORR) of 15%. However, gefitinib was removed from the market in 2005 due to perceived ineffectiveness at the time with exceptions granted to patients already receiving and benefiting from gefitinib or for use in clinical trials. In 2015, gefitinib returned to the U.S. market with FDA approval for first-line therapy in patients with metastatic NSCLC with confirmed EGFR mutations (EGFR exon 19 deletions or exon 21 L858R substitution) based on biomarker testing. The new approval was based on a multicenter study of treatment-naive patients with metastatic EGFR-positive NSCLC. An ORR of 50% was found, with median duration of response of 6 months.
• Most Common Adverse Events: diarrhea, rash, acne, dry and itchy skin, decreased appetite, shortness of breath

Afatinib (Gilotrif, Boehringer Ingelheim):

• Second-generation irreversible EGFR TKI approved by the FDA in 2013 for first-line therapy of patients with commonEGFR-mutated metastatic NSCLC. The approval was based on an international, multicenter phase 3 clinical trial of patients with metastatic EGFR-mutated NSCLC. An ORR of 50.4% was found, with median PFS of 11.1 months. In 2018, the FDA broadened approval to the first-line setting for patients with metastatic NSCLC positive for other nonresistant EGFR mutations (including S768I, L861Q, and/or G719X). This indication was based on a subset of patients from LUX-LUNG 2, 3, and 6 clinical trials. An ORR of 66% was found across these three trials, with 52% of patients experiencing a duration of response of 12 months or longer.
• Most Common Adverse Events: diarrhea, rash, dry skin, decreased appetite, vomiting, fatigue, sore mouth, skin infection around the nails, itchy skin,shortness of breath

Osimertinib (Tagrisso, AstraZeneca):

• Third-generation EGFR TKI was granted accelerated approval by the FDA in 2015 with full approval in 2017 for the treatment of patients with metastaticEGFR T790M mutation-positive NSCLC whose disease progressed after EGFR TKI therapy. In 2018, osimertinib was granted broadened approval by the FDA for first-line therapy of patients with metastatic NSCLC with common EGFR mutations. This approval was based on the international, multicenter FLAURA clinical trial of unresectable or metastatic NSCLC patients with EGFR exon 19 deletions or exon 21 L858R substitution who progressed on prior systemic therapy. At the time of approval, the median PFS was 18.9 months vs. 10.2 months with first-generation EGFR TKIs (gefitinib or erlotinib). Updated results from the phase 3 FLAURA trial showed median OS of 38.6 months compared with 31.8 months with erlotinib or gefitinib and enhanced central nervous system (CNS) penetration and efficacy (PFS of 15.2 months vs. 9.6 months, respectively).
• Most Common Adverse Events: diarrhea, nausea, rash, dry skin, nail toxicity, sores mouth, decreased appetite

Dacomitinib (Vizimpro, Pfizer):

• Second-generation irreversible EGFR TKI approved by the FDA in 2018 for first-line therapy of patients with metastatic NSCLC withEGFR exon 19 deletions or exon 21 L858R substitution mutations. The approval was based on the multicenter ARCHER 1050 clinical trial of patients with unresectable, metastatic NSCLC with common EGFR mutations and a minimum of 12 months disease-free post-systemic therapy. No significant difference was found in ORR and OS between patients treated with dacomitinib or gefitinib; however, a significant improvement in PFS of 14.7 months vs. 9.2 months, respectively, was found.
• Most Common Adverse Events: diarrhea, interstitial lung disease, rash, skin infection around the nails, sore mouth, decreased appetite, dry skin, decreased weight, cough, itchy and dry skin, hair loss

EGFR Exon 20 Insertions

Background

The third most common type of EGFR mutations seen in patients with NSCLC are exon 20 insertions, accounting for 4-12% of EGFR and 1-2% of all NSCLC mutations. Exon 20 insertions are considered particularly difficult to treat due to their poor response to standard first-, second-, and third- generation EGFR TKIs. Efficacy of current targeted therapies is dramatically impacted by which amino acids are affected by the 3-21 base pair insertion in EGFR exon 20. Due to poor targeted therapy response rates, the presence of EGFR exon 20 insertions are associated with worse patient outcomes. 

Clinical Features Associated

Females, minimal/no smoking history, Asian ethnicity, adenocarcinoma histology, aggressive tumors, bone and CNS metastasis

Detection Methods

NGS; PCR should be considered carefully as it has been shown to miss approximately 50% of exon 20 insertions.

Most Common Alterations

• Insertions: Representation of EGFR exon 20 mutations are shown in figure below.   
  • Of note: While most insertions identified to date do not respond to EGFR TKIs, exon 20 A763_Y764insFQEA has shown sensitivity to first-generation EGFR TKI erlotinib.

Acquired Resistance Mutations

The most commonly acquired resistance mutation is T790M, methionine substituted for threonine, within the active site of EGFR, accounting for more than half of all acquired resistance mutations from first-generation EGFR inhibitors in patients with NSCLC. 

FDA-Approved Targeted Therapies:

Osimertinib (Tagrisso, AstraZeneca):

• Third-generation EGFR TKI granted accelerated approval from the FDA in 2015 with full approval in 2017 for the treatment of patients with metastaticEGFR T790M-positive NSCLC who progressed on or after EGFR TKI therapy. Accelerated approval was granted based on two single-arm clinical trials — AURA and AURA2 — that showed a combined ORR of 59%. Full approval was based on the multicenter AURA3 clinical trial of patients with metastatic EGFR T790M-positive NSCLC who progressed on first-line EGFR TKI therapy. PFS was significantly improved in patients treated with osimertinib (10.1 months) compared with those treated with chemotherapy (4.4 months). In addition, an ORR of 57% was found in those patients with CNS tumor.
• Most Common Adverse Events: diarrhea, rash, dry skin, nail toxicity, fatigue, low white blood cell count

Amivantamab (Rybrevant, Janssen):

• Monoclonal antibody therapy approved by the FDA in 2021 as front-line therapy for adult patients with NSCLC with EGFR exon 20 insertion mutations. The approval was based on the CHRYSALIS study of patients with EGFR 20 insertion-mutated NSCLC whose disease progressed on or after chemotherapy. At the time of approval, an ORR of 40% was found with a median duration of response of 11.1 months; 63% of patients experienced a duration of response of 6 months or longer. 
• Most Common Adverse Events: rash, infusion-related reactions, skin infections around the nails, muscle and joint pain, shortness of breath, nausea, fatigue, swelling in the extremities or face, mouth sores, cough, constipation, vomiting, altered blood tests

Mobocertinib (Exkivity, Takeda):

• An irreversible, small-molecule TKI granted approval by the FDA in 2021 for adults with locally advanced or metastatic NSCLC withEGFR exon 20 insertion mutations. The approval was based on an international, multi-cohort clinical trial, Study 101, of patients with locally advanced or metastatic NSCLC with EGFR exon 20 insertion mutations. At the time of approval, an ORR of 28% was found with a median duration of response of 17.5 months.
• Most Common Adverse Events: diarrhea, rash, nausea, vomiting, decreased appetite, fatigue, dry skin, sore mouth, skin infections around the nails, muscle pain

EGFR Exon 18

Background

The rare EGFR mutations that occur in exon 18 involve missense mutations (G719X and E709X), insertion-deletion mutations (ie, E709_T710delinsX), and other molecular subtypes. In total, these mutations account for approximately 3-5% of EGFR alterations in NSCLC. Little is currently known about exon 18 mutations, with most literature based on small retrospective studies. These alterations do not to respond well to first-generation EGFR TKIs.

Clinical Features Associated

Smoking/no smoking history, no sex predilection, adenocarcinoma histology

Detection Methods

PCR, NGS

Most Common Alterations

The most common exon 18 mutation is G719X (A, C, D, S, or V), followed by E709X, E709_T710delinsX, and finally other subtypes. Studies have shown the best response with second-generation EGFR TKIs. G719X has shown the greatest response to afatinib and neratinib (Nerlynx, Puma Biotechnology) with ORRs of 77.8% and 75%, respectively.

FDA-Approved Targeted Therapy

Afatinib (Gilotrif, Boehringer Ingelheim):

• Second-generation irreversible EGFR TKI granted broadened approval by the FDA in the first-line setting for patients with metastatic NSCLC positive for uncommon EGFR mutations (including S768I, L861Q and/or exon 18 G719X). The approval was based on a subset of patients from LUX-LUNG 2, 3, and 6 clinical trials. An ORR of 66% was found with 52% of patients experiencing duration of response of 12 months or longer and an ORR of 77.8% in patients with exon 18 G719X mutation.
• Most Common Adverse Events: diarrhea, rash, dry skin, decreased appetite, vomiting, fatigue, sore mouth, skin infection around the nails, itchy skin, shortness of breath

ALK: Anaplastic Lymphoma Kinase

Background

A transmembrane tyrosine kinase, belonging to the insulin receptor superfamily, found across several tumor types. While more commonly found in NSCLC (3-7%), ALK-positive mutations have also been found in a small number of patients with small-cell lung cancer. Three types of ALK mutations have been identified in NSCLC — rearrangement, amplification and point mutations — with the most common being ALK rearrangements. ALK rearrangements, often in combination with EGFR mutations or ROS1 rearrangements, lead to activation of various signaling pathways associated with cell proliferation and survival. Similar to what is seen in EGFR, ALK mutations can form as a secondary resistance to treatment with ALK-TKIs.

Clinical Features Associated

Younger age, minimal/no smoking history, adenocarcinoma histology

Detection Methods

FISH, IHC, NGS

Most Common Alterations

• ALK Rearrangements involve a portion of the ALK gene breaking and attaching to another gene to form fusion oncogene. ALK rearrangements exhibit lower response rates to immunotherapy, specifically immune checkpoint inhibitors. Due to the approval of targeted therapies, 5-year OS has increased for patients with NSCLC with ALK rearrangements.
• EML4-ALK Fusion Oncogene (Echinoderm microtubule-associated protein like-4) was the first and major ALK-fusion partner identified. This inversion rearrangement involving a portion of EML4 gene fusing with a portion of the ALK gene leads to constitutively active ALK kinase resulting in malignant growth and proliferation. Multiple variants of this fusion oncogene have been identified and are found in 3%-7% of patients with NSCLC.
  • Over 90 ALK-fusion partners have been reported in NSCLC, the majority of which have been identified from 2018 onward due, in part, to the adoption of NGS for molecular profiling. 

•  Secondary Acquired Resistance Mutations: Secondary ALK kinase domain resistance mutations have been identified after progression on ALK-TKIs.
  • Crizotinib resistance is commonly associated with the following secondary ALK mutations: G1269A, gatekeep mutation L1196M, T1151R, C1156Y, R1192P, G1202R, S1206Y, A1280V, and L1535Q.

FDA-Approved Therapies

Targeted therapy is the preferred first-line treatment and is not recommended in combination with immunotherapy.

Crizotinib (Xalkori; Pfizer, EMD Serono):

• First-generation multi-targeted ATP-competitive TKI that confers activity againstMET, ALK and ROS1. FDA granted accelerated approval in 2011, with full approval granted in 2013 for patients with metastatic NSCLC positive for ALK rearrangements. Approval was based on the PROFILE 1014 study. At time of approval, an overall response rate (ORR) of 74% was found with median PFS of over 10 months. Of note, nearly all patients treated with front-line crizontinib eventually progress and develop resistance mechanisms.
• Most Common Adverse Events: vision disorders, nausea, diarrhea, vomiting, swelling, constipation, elevated liver enzymes, fatigue, decreased appetite, upper respiratory infection, dizziness/numbness/tingling in the hands and/or feet

Ceritinib (Zykadia, Novartis):

• Second-generation multi-targeted, brain-penetrable inhibitor againstALK and ROS1 was granted approval by FDA in 2014 as second-line therapy for patients with metastatic ALK-fusion-positive NSCLC post-crizotinib. In 2017, the FDA broadened approval to the first-line setting for these patients based on the ASCEND-4 study of treatment-naive, metastatic ALK-positive NSCLC with stable brain metastasis. Median PFS of 16.6 months was found with ceritinib vs. 8.1 months in the chemotherapy cohort. The ASCEND-8 trial evaluated a lower dose of ceritinib, with median PFS of 17.6 months compared with 10.9 months in the higher dose cohort and reduction in adverse events reported.
• Most Common Adverse Events: diarrhea, nausea, vomiting, fatigue, abdominal pain, decreased appetite, cough — Of note, serious adverse reactions occurred in 38% of patients treated with ceritinib in the above trial.

Alectinib (Alecensa, Genentech):

• Second-generation potent, brain-penetrable ALK inhibitor was granted approval by FDA in 2015 as second-line therapy for patients with metastaticALK-fusion-positive NSCLC post-crizotinib. In 2017, the FDA expanded approval to first-line setting for these patients. The approval was based on the ALEX study of treatment-naive patients with advanced/metastatic ALK-positive NSCLC. At the time of approval, a PFS of 34.8 months was found with alectinib vs. 10.9 months with crizotinib. Alectinib has shown superior efficacy duration with low incidence of serious adverse events after long-term use compared with crizotinib. Due to alectinib’s blood-brain barrier permeability, studies have shown enhanced prevention of brain metastases and longer PFS.
• Most Common Adverse Events: fatigue, constipation, swelling, muscle pain, anemia — Of note, serious adverse reactions occurred in 28% of patients treated with alectinib in the above study.

Brigatinib (Alunbrig, Takeda):

• Second-generation, brain-penetrable, multitargeted TKI against ALK, mutant EGFR, and ROS1 was approved by the FDA in 2017 as second-line therapy for patients with metastatic ALK-fusion-positive NSCLC after crizotinib. In 2020, the FDA broadened approval to the front-line setting based on the ALTA-1L study. At the time of approval, a median PFS of 24 months was found in patients treated with brigatinib compared with 11 months in those treated with crizotinib and an ORR of 74% compared with 62%, respectively.
• Most Common Adverse Events: diarrhea, fatigue, nausea, rash, cough, myalgia, headache, hypertension, vomiting, shortness of breath

Lorlatinib (Lorbrena, Pfizer): 

• Third-generation multi-kinase inhibitor against ALK and ROS1 was approved by FDA in 2018 for patients with metastatic ALK-positive NSCLC for patients previously treated on one or more ALK-TKIs. Lorlatinib was designed specifically against ALK drug resistance mutation sites and with high blood-brain permeability. In 2021, full approval was granted by the FDA in the front-line setting based on the CROWN study of treatment-naive, ALK-positive stage IIIB/IV NSCLC. At the time of approval, an ORR of 76% was found with lorlatinib compared with 58% with crizotinib. Median PFS and OS data were still immature, but initial results suggested improvement with lorlatinib compared with crizotinib.
• Most Common Adverse Events: grade 3 laboratory abnormalities, swelling, dizziness/numbness/tingling in the hands or feet, weight gain, cognitive effects, fatigue, shortness of breath, joint paint, diarrhea, mood effects, high blood cholesterol and triglycerides, cough — Of note, serious adverse events that required discontinuation of therapy were 7% and 9% in lorlatinib and crizotinib, respectively, in the above trial.

ROS1: ROS Proto-oncogene 1

Background

A transmembrane tyrosine kinase gene of the insulin receptor family involved in cell proliferation. ROS1 rearrangements were first identified in patients with NSCLC in 2007 and account for 1% to 2% of NSCLC driver mutations. These mutations are the result of rearrangements of ROS1 with other genes forming fusion proteins. The most common fusion is with CD74. ROS1 fusion proteins result in constitutive ROS1 kinase activity leading to upregulation of downstream signaling pathways including MAPK, PI3K/AKT and JAK pathways known to play a role in malignant transformation. While most fusions are mutually exclusive of other oncogenic drivers, there are some instances of concurrent EGFR, MET BRAF, and KRAS mutations. ROS1 rearrangements are typically associated with a better prognosis compared with other driver mutations.

Clinical Features Associated

Younger age, minimal/no smoking history, females, adenocarcinoma histology

Detection Methods

FISH, IHC (in combination with another detection method), RT-PCR, NGS

Most Common Alterations

• Fusion Partners: The most common identified fusion partners include: CD74, SLC34A2, SDC4, EZR, FIG, TPM3, LRIG3, KDELR2, CCDC6, MSN, TMEM106B, TPD52L, CLTC, WNK1, MYO5C, TFG, RBPMS and LIMA1
• CD74: Most common (40% to 45%) and associated with increased risk of brain metastases and lower ORR to crizotinib.

Secondary Acquired Resistance Mutations

Acquired resistance to TKI therapy is an issue in patients with ROS1 rearrangements resulting in secondary mutations, including: G2032R, D2033N, S1986Y/F, L2026M and L1951R.  The most frequent secondary mutation is G2032R.

FDA Approved Targeted Therapies

Crizotinib (Xalkori; Pfizer, EMD Serono):

• First-generation, multi-targeted ATP-competitive TKI that confers activity againstMET, ALK and ROS1 with poor blood-brain barrier penetration. FDA approval was granted in 2016 for the treatment of patients with metastatic ROS1-positive NSCLC based on the phase 1 PROFILE 1001 study of ROS1-rearranged NSCLC patients. At time of approval, an ORR of 72% was found, with a disease control rate of 90% and median PFS of 19.2 months. In 2017 NCCN guidelines recommended crizotinib for patients with NSCLC with ROS1 rearrangements. Despite initial positive responses, most patients treated with crizotinib eventually progress and often develop brain metastasis.
• Most Common Adverse Events: vision disorders, nausea, diarrhea, vomiting, swelling, constipation, elevated liver enzymes, fatigue, decreased appetite, upper respiratory infection, dizziness/numbness/tingling in the hands and/or feet

Entrectinib (Rozlytrek, Genentech):

• Second-generation multi-kinase ATP-competitive inhibitor granted approval by the FDA in 2019 for patients with metastatic ROS1-positive rearranged NSCLC based on the integrated analysis of phase 1 STARTRK-1, phase 2 STARTRK-2, and phase 2 ALKA-372-001 trials. Combined results at the time of approval showed an ORR of 77.4% with a median duration of response of 24.6 months. Entrectinib is recommended over crizotinib for patients with CNS involvement due to the enhanced intracranial penetration. Entrectinib has not been shown effective in the following ROS1 mutations: L2026M, G2032R and D2033N.
• Most Common Adverse Events: fatigue, constipation, altered taste, swelling, dizziness, diarrhea, nausea, abnormal sensation, shortness of breath, muscle pain, cognitive impairment, increased weight, cough, vomiting, fever, joint stiffness, vision disorders

BRAF: v-raf murine sarcoma viral oncogene homolog B

Background

A serine/threonine kinase in the RAS/mitogen-activated protein kinase (MAPK) signaling pathway. Alterations in BRAF result in continuous activation of the MAPK pathway, promoting cell growth, proliferation and differentiation. BRAF mutations occur in 2% to 4% of patients with NSCLC, with about half occurring in the V600 position where a glutamic acid substitution is found (V600E). While non-V600E may coexist with KRAS mutations, other BRAF mutations are mutually exclusive from EGFR mutations and ALK rearrangements in treatment-naive patients.     

Clinical Features Associated

Smoking history, Caucasians, females, adenocarcinoma histology

Detection Methods

RT-PCR, NGS, and Sanger sequencing. Liquid biopsy is an option for patients who cannot undergo biopsy or do not have enough tissue remaining for NGS.

Classes of BRAF Mutations

Class I:

• Most frequently identified, RAS-independent, strong activation of BRAF kinase activity, and continuous activation of MAPK pathway.
• V600E: 1-2% of lung adenocarcinomas and 50% of BRAF-mutated NSCLC. BRAF V600E mutation occurs mutually exclusive of KRAS, EGFR and ALK alterations and is currently the only BRAF mutation with FDA-approved targeted therapy. Combination BRAF-MEK inhibitors have shown better clinical outcomes than platinum-based chemotherapy; however, acquired resistance to this combination remains a challenge.
  • Clinical Features Associated: females, not correlated with smoking history

Class II:

• RAS-independent, including both high and intermediate BRAF kinase activity based on MAPK pathway activation, and constitutively active dimers.
• The most common is G469A (35% of all BRAF mutations).
• Other identified mutations include K601E and L597Q.

Class III:

• RAS-dependent, BRAF kinase activity is low or impaired, requires upstream activation of MAPK pathway and often coexists with RAS mutations or NF1 loss.
• Most common: D594X (10% of all BRAF mutations)
• Other identified mutations include G466X.

Additional missense, deletions and mutations of unknown function have also been identified

FDA-Approved Targeted Therapy

Dabrafenib Plus Trametinib (Novartis):

• This BRAF + MEK combination therapy was granted full approval by the FDA in 2017 and is the only approved treatment for patients with metastatic NSCLC with BRAF V600E mutation. The approval was based on the BRF113928 multicenter study of patients with locally confirmed BRAF V600E mutation-positive NSCLC. At time of approval, an ORR of 63% and duration of response of 12.6 months were found in patients who received prior lines of chemotherapy. For treatment-naive patients, an ORR of 61% was found, with 59% of responders having response of greater than 6 months. 
• Most Common Adverse Events: fever, fatigue, nausea, vomiting, diarrhea, dry skin, decreased appetite, swelling, rash, chills, bleeding, cough, shortness of breath

NTRK: Fusion Neurotrophic Tropomyosin Receptor Kinase

Background

NTRK genes encode tropomyosin receptor kinases (TRKA/B/C) involved in development and function of the nervous system. While NTRK amplifications and mutations occur, they are not strong drivers of cancer development associated with NTRK fusions. Fusions form when a portion of the NTRK gene (3’ end) fuses with a portion of another gene (5’ end), resulting in continual activation of various signal transduction pathways leading to cancer cell proliferation, transformation and invasiveness. NTRK fusions have been identified in less than 1% of patients with NSCLC and tend to be mutually exclusive of other mutational drivers in treatment-naive patients.

Clinical Features Associated

Some studies have shown association with younger age, minimal/no smoking history, multiple sites of metastasis, squamous and non-squamous with majority adenocarcinoma histology.

Detection Methods

NGS (preferred method), RT-PCR, FISH, IHC

• Fusion: IHC, FISH, RT-PCR, NGS
• Amplification: IHC, FISH, NGS
• Mutation: NGS

Most Common Alterations

NTRK Fusion Genes (NTRK1/2/3): Majority fusions identified in NSCLC are NTRK1 and NTRK3.

NTRK Secondary Acquired Resistance Mutations

Develop after first-generation TKI therapy for NTRK fusion. Multiple mechanisms of resistance have been identified, with gene rearrangements being the most common, often resulting in substitution of a larger amino acid for a smaller amino acid in the ATP binding site of the TKI drug — ie, TRKA G595R and G667C and TRKC G623R and G696A. This substitution interferes with the ability of the TKI to bind and interrupt the oncogenic pathway. Next-generation drugs were developed, including selitrectinib (selective TRK inhibitor, Bayer) and repotrectinib (TRK/ROS1 inhibitor, Turning Point Therapeutics) that are smaller than first-generation TKIs and are able to shut down oncogenic signaling again.

FDA-Approved Targeted Therapies

Larotrectinib (Vitrakvi, Bayer):

• First-generation highly selective inhibitor of all three TRK kinases was granted approval by the FDA in 2018 for adult and pediatric patients with solid tumors withNTRK gene fusions without known acquired resistance mutation, metastatic or no other viable effective treatments. Larotrectinib has been shown to cross the blood-brain barrier. Approval was based on the results of three multicenter clinical trials (LOXO-TRK-14001, SCOUT and NAVIGATE). Combined results at time of approval showed an ORR of 75%, including 22% patients with complete responses and 53% patients with partial responses; 63% of patients experienced a duration of response of 9 months or longer at time of data lock. A separate analysis of patients with NTRK fusion-positive lung cancer found an ORR of 73%, median duration of response (DoR) of 33.9 months, PFS of 35.4 months and OS of 40.7 months.
• Most Common Adverse Events: fatigue, nausea, dizziness, vomiting, cough, elevated liver enzymes (AST), constipation, diarrhea

Entrectinib (Rozlytrek, Genentech):

• First-generation TRK inhibitor against all three TRKs, ROS1 and ALK was granted approval by the FDA in 2019 for patients 12 years of age and older withNTRK fusion-positive solid tumors with no known acquired resistance mutations, metastatic and no available effective treatment options. Entrectinib is able to cross the blood-brain barrier but is not effective against NTRK secondary mutations. Approval was based on data from multiple clinical trials of NTRK fusion-positive patients (ALKA, STARTRK-1, STARTRK-2, STARTRK-NG). A combined ORR of 57% was found with 68% of patients exhibiting response for 6 months or longer at time of approval. An analysis of NTRK fusion-positive NSCLC patients found an ORR of 67% with a median PFS and OS of 14.9 months.
• Most Common Adverse Events: fatigue, constipation, altered taste, swelling, dizziness, diarrhea, nausea, trouble breathing, impairment of sensitivity often to touch, muscle and joint pain, cognitive impairment, increased weight, cough, vomiting, vision disorders, fever

Repotrectinib:

• Second-generation multi-kinase inhibitor against all three TRKs, ROS1 and ALK was granted breakthrough therapy designation by the FDA in fall of 2021 for patients with advanced solid tumors with NTRK gene fusion who progressed on prior TRK inhibitors with or without chemotherapy and no alternative options available. Repotrectinib is able to cross the blood-brain barrier. The approval was based on preliminary data from the phase 1/2 TRIDENT-1 study of patients with NTRK fusion-positive NSCLC. An ORR of 50% in six patients with TKI-pretreated advanced NTRK-positive solid tumors.
• Most Common Adverse Events: dizziness, fatigue, constipation, trouble breathing, altered taste

MET: Mesenchymal-to-Epithelial Transition

Background

A TKI for hepatocyte growth factor (HGF). Overexpression, gene amplification, rearrangements, or mutations of MET protein result in alterations to MET signaling pathway, leading to downstream effects on cell growth, apoptosis, motility and invasiveness.

Detection Methods

• MET exon 14 skipping: recommend RNA NGS, but qRT-PCR or DNA NGS are other options.
• MET amplification: NGS or FISH. Note: If a lung biopsy is not performed or not enough tissue exists to perform NGS or FISH, liquid biopsy can be used to identify markers in the blood.

Most Common Alterations

• MET Mutation: MET mutations, specifically MET exon 14-skipping, occur in 2% to 4% of NSCLC. MET exon 14-skipping mutations result in reduced degradation of MET protein causing MET to behave in an oncogenic manner. They have been shown to be a highly aggressive subtype with poor prognosis. Current research suggests these mutations are independent drivers and are mutually exclusive of other known driver mutations. Hundreds of alterations have been identified, including point mutations, deletions, insertions or complex mutations resulting in exon 14-skipping. Loss of exon 14 due to a splicing event results in increased stability of the receptor, which leads to increased kinase activity and tumor growth.
  • Clinical Features Associated: older age, smoking and no smoking history, pulmonary sarcomatoid carcinoma (PSC), adenocarcinoma histology, adenosquamous carcinoma
• MET Rearrangement/Fusion: MET fusions were first discovered in cell lines exposed to carcinogen. The true rate in NSCLC, based on NGS, is less than 1% of identified fusions effect the function of the gene in oncogenic manner.
  • Includes: TPR, TRIM4, ZKSCAN1, PPFIB1, LRRFIP1, EPS15, DCTN1, PTPRZ1, NTRK1, CLIP2, TFG, HLA-DRB1, KIF5B
• MET Amplification: MET amplification can be a primary event, occurring in 2% to 5% of newly diagnosed patients with NSCLC, or a secondary event, associated with acquired resistance to TKIs, primarily EGFR TKIs (50% to 60% of first- and second-generation TKI acquired resistance). High MET amplification is very rare (<0.34%).

FDA-Approved Targeted Therapy

Crizotinib (Xalkori; Pfizer, EMD Serono):

• A multi-kinase, ATP-competitive TKI that confers activity againstMET, ALK and ROS1. This type 1 MET inhibitor received breakthrough approval by the FDA in 2018 for patients with MET exon 14 altered NSCLC who progressed on platinum-based chemotherapy. The approval was based on an expansion cohort of the PROFILE 1001 study of patients with MET exon 14-altered NSCLC treated with crizotinib. In 18 evaluable patients, a partial response of 44% was observed, with 50% experiencing stable disease at time of analysis.
• Most Common Adverse Events: diarrhea, abdominal pain, swelling, nausea, vision disorders, decreased heart rate, difficulty breathing, vomiting

Capmatinib (Tabrecta, Novartis):

• Type 1 MET inhibitor received approval by the FDA in 2020 for patients with metastaticMET exon 14 skipping mutated NSCLC in front-line and refractory setting. Approval was based on the GEOMETRY mono-1 trial of patients with metastatic NSCLC confirmed MET exon 14 skipping. At time of approval, an ORR of 68% was found among treatment-naive patients with a DoR of 12.6 months. An ORR of 41% was found among previously treated patients with DoR 9.7 months.
• Most Common Adverse Events: lower limb swelling, nausea, fatigue, vomiting, difficulty breathing, decreased appetite — Of note additional adverse events include interstitial lung disease, hepatotoxicity, photosensitivity, and embryo-fetal toxicity.

Tepotinib (Tepmetko, Merck/EMD Serono):

• Type 1 MET inhibitor granted accelerated approval by the FDA in early 2021 for patients with metastatic NSCLC with MET exon 14 skipping mutation. It is a highly selective, ATP-competitive, reversible MET TKI that may penetrate the CNS. The approval was based on the VISION trial of advanced or patients with metastatic NSCLC with MET exon 14 skipping alterations. At time of approval, an ORR of 43% was found for treatment-naive and previously treated patients with a median DoR of 10.8 months and 11.1 months, respectively.
• Most Common Adverse Events: swelling, fatigue, nausea, diarrhea, musculoskeletal pain, difficulty breathing — Of note, additional adverse events include interstitial lung disease, hepatotoxicity and embryo-fetal toxicity

RET Fusion: Rearranged During Transfection

Background

A TKI involved in cell proliferation, migration and differentiation. RET chromosomal rearrangements, the product of intact tyrosine kinase domain of RET protein fusing with a portion of another gene, were first identified in lung adenocarcinoma in 2012. These fusions result in constitutive activation of RET, which promotes cancer cell growth, proliferation and survival. RET fusions have been identified in 1% to 2% of NSCLC and are exclusive of EGFR, ALK, KRAS and BRAF mutations.

Clinical Features Associated

Younger age, minimal/no smoking history, adenocarcinoma histology — the majority of patients have stage IV disease and have early lymph node metastasis and higher risk of brain metastasis.

Detection Methods

FISH, IHC, RT-PCR, NGS, liquid biopsy

Most Common Alterations

• Fusion Partners: The most common and well characterized of RET fusion partners is KIF5B, accounting for approximately 70% of RET rearrangements followed by CCDC6 (10-25%) and NCOA4. 
• Additional Fusion Partners Identified Include: CUX1, MYO5C, EPHA5, TRIM33, CLIP1, ERC1, PICALM, FRMD4A, RUFY2, KIAA1468, KIAA1217, ZNF477P, HTR4, and TRIM24.
• Acquired Resistance Mutations: The most common mutations identified post-TKI include RET V804 gatekeeper mutation, G810S/R/C, and S904F. 

FDA-Approved Targeted Therapies

Selpercatinib (Retevmo, Eli Lilly):

• Highly selective ATP-competitive small-molecule RET inhibitor received approval by the FDA in 2020 for the treatment of advanced RET-rearranged NSCLC. Selpercatinib has been shown to have significant CNS penetration and low drug interactions. Approval was based on the phase 1/2 LIBRETTO-001 study of patients with advanced RET fusion-positive NSCLC. At time of approval, an ORR of 68% was found in previously treated patients with a median PFS of 18.4 months. An ORR of 85% was found in treatment-naive patients. Updated results of the LIBRETTO-001 study in 2019 showed an ORR in patients with CNS metastasis of 91%. 
• Most Common Adverse Events: dry mouth, diarrhea, hypertension, elevated liver enzymes, fatigue, constipation, headache, nausea, elevated creatine levels, swelling of the hands and legs

Pralsetinib (Gavreto; Blueprint Medicines, Genentech):

• Highly selective RET inhibitor was granted approval by the FDA in 2020 for adults with metastatic RET fusion-positive NSCLC. Pralsetinib has been shown effective in both patients with NSCLC with primaryRET fusions and mutations as well as acquired RET resistance mutations. Approval was based on the phase 1/2 ARROW multi-cohort trial of patients with RET-positive NSCLC. At the time of approval, an ORR of 57% was found for previously treated patients with 5.7% experiencing a complete response and 80% experiencing a response of 6 months or longer. An ORR of 70% was found in treatment-naive patients, with 11% experiencing a complete response and 58% experiencing a response of 6 months or longer. Additionally, an ORR of 55% was found in patients with measurable brain metastasis
• Most Common Adverse Events: fatigue, elevated liver enzymes, anemia, constipation, hypertension, low white blood cell count, diarrhea, elevated creatine, low neutrophil count, musculoskeletal pain

HER2: Human Epidermal Growth Factor 2

Background

HER2 is part of the ERBB receptor tyrosine kinase family. Also known as ERBB2, HER2 does not have its own ligand binding domain, therefore it cannot bind growth factors. Alterations in the HER2 gene lead to hyperactivation triggering multiple signaling pathways, including MAPK and PI3K/AKT, resulting in uncontrolled cell proliferation, transformation and oncogenesis. HER2 alterations occur through three potential mechanisms: gene mutation, amplification and protein overexpression. HER2 amplifications occur in less than 15% patients with NSCLC. HER2 mutations are seen in 2% to 6% of EGFR/KRAS/ALK-negative NSCLC. While targeted therapies have been successful in breast cancer, they have not proven effective in NSCLC.

Clinical Features Associated

Females, Asian ethnicity, minimal/no smoking history, adenocarcinoma histology, CNS involvement reported in up to 47% of patients

Detection Methods

FISH, IHC, RT-PCR, NGS

Most Common Alterations

HER2 Mutations:

• HER2 mutations are found in 1% to 4% of patients with NSCLC. The most common mutations occur in exon 20 which resemble EGFR 20-activating mutations
• YMVA exon 20 mutations, insertion of three to 12 base pairs resulting in duplication or insertion of amino acids, account for 80% to 90% of all HER2 mutations and are the focus of HER2-directed therapies.
• Point mutations in exon 20, L755S and G776C, account for 8% to 10% of HER2 mutations.
• Mutations found in conjunction with other driver mutations, including EGFR, ALK and ROS, should be treated differently. Additional rare mutations have been identified (G660D, R678Q, E693 and Q709L) but their prognostic value remains unclear.
• Current Diagnostic Methods: NGS, RT-PCR, and qPCR

HER2 Gene Amplification:

• Amplifications have been identified in 2% to 5% of treatment-naive NSCLC patients and 13% in those patients who progressed on prior EGFR TKI therapy. Prognosis for patients with HER2 amplification remains unclear, with some studies suggesting shorter OS
• Current Diagnostic Methods: FISH, NGS, ELISA, and RT-PCR

HER2 Overexpression:

• Overexpression is found in 2% to 30% of patients with NSCLC and has been shown to be associated with worse survival especially in early-stage NSCLC
• Current Diagnostic Methods: IHC, qPCR and RT-PCR

FDA-Approved Targeted Therapies

Trastuzumab deruxtecan (Enhertu; AstraZeneca, Daiichi Sankyo):

• HER2-directed antibody drug-conjugate received FDA approval in 2020 for patients with HER2 mutant metastatic NSCLC who have progressed on platinum-based chemotherapy treatment. Approval was based on the DESTINY-Lung01 multicenter phase 2 study of patients with HER2-mutant or HER2 overexpression unresectable and metastatic NSCLC. At the time of approval, an ORR of 72.7% was found with a median PFS of 11.3 months
• Most Common Adverse Events: nausea, anemia, decreased appetite, decreased neutrophil, hair loss

Poziotinib (Spectrum Pharmaceuticals):

• Irreversible TKI targets EGFR and HER2 exon 20 mutations received fast-track designation from the FDA in 2021 for patients with NSCLC with HER2 exon 20 mutations. Designation was granted based on two studies. Single-centered phase 2 trial of heavily pretreated NSCLC patients with either EGFR or HER2 mutations found an ORR of 42% with median PFS of 5.6 months in the HER2 mutant cohort. The single center ZENITH20 trial included patients with NSCLC with mutations in exon 20 in either EGFR or HER2. At a median follow up of 8.3 months, an ORR of 27.8% was found with a disease control rate of 70% and median PFS of 5.5 months
• Most Common Adverse Events: rash, diarrhea, inflamed or sore mouth

KRAS: Kirsten RAT Sarcoma Viral Oncogenic Homolog

Background

KRAS is the most frequently mutated oncogene in human cancers and is often associated with resistance to targeted therapies and poor outcomes. KRAS acts like a switch to activate downstream signaling pathways, including MAPK and PI3K, involved in cell proliferation, cell cycle regulation, metabolic changes, cell survival and cell differentiation. Dysregulated KRAS has been associated with tumor growth, cancer cell survival, invasion and migration. Alterations in KRAS account for approximately 25% of mutations in NSCLC and are found exclusive of EGFR and ALK mutations in treatment-naive patients.

Clinical Features Associated

Caucasian or African ancestry, males, smoking history, history of exposure asbestos, adenocarcinoma histology

Most Common Alterations

• Point Mutations: The most common point mutations are identified in codons 12 and 13 and include G12C, G12V, and G12D. Conflicting data exists in the literature regarding the prognostic value of mutational subtypes, with some finding worse prognosis associated with G12C and G12V and others finding no differences.
  • The most common point mutation is KRAS G12C, accounting for around 39% of KRAS mutations and 12% to 13% of all mutations identified in NSCLC. G12C is often associated with smokers, while G12D is more common in nonsmokers.

• Co-mutational Partners: The most common mutational partners include TP53 (40%), STK11/LKB1 (32%), and CDKN2A (19.8%). The presence of these co-mutational partners has been associated with altered response rates to therapy. STK11/LKB1 has been linked to greater resistance to immune checkpoint inhibitors in NSCLC. TP53/KRAS co-mutation is associated with enhanced PD-L1 expression; therefore, patients have seen clinical benefit from PD-1 inhibitors. Of note: STK11/LKB1 and CDKN2A have been associated with worse prognosis compared with TP53

FDA-Approved Targeted Therapies

Sotorasib (Lumakras, Amgen):

• Irreversible RAS GTPase inhibitor received accelerated approval by the FDA in 2021 for adult patients withKRAS G12C mutated locally advanced or metastatic NSCLC and at least one prior line of systemic therapy. Approval was based on the CodeBreak 100, multicenter clinical trial of patients with locally advanced or metastatic KRAS G12C mutated NSCLC. An ORR of 36% was found with a median duration of response at time of approval of 10 months 
• Most common adverse events: diarrhea, joint and muscle pain, nausea, fatigue, liver damage, cough

Adagrasib (MRTX849, Mirati Therapeutics Inc.):

• Highly selective inhibitor ofKRAS G12C received breakthrough designation by the FDA in 2021 for patients with KRAS G12C-mutated locally advanced or metastatic NSCLC and at least one prior line of systemic therapy. This designation was based on the preliminary results from the phase 1/2 KRYSTAL-01 clinical trial of patients with advanced NSCLC who harbor the KRAS G12C mutation and progressed on prior systemic therapy. An ORR of 64% with 45% of patients experiencing a partial response was found at time of designation.
• Most common adverse events: diarrhea, joint and muscle pain, nausea, fatigue, liver damage, cough