Despite Progress, More Actionable Mutations Needed in Lung Cancer

Lung cancer has emerged as a prime example of both the promise and pitfalls of precision medicine. Significant progress has been made in the treatment of patients whose tumors have specific molecular drivers, such as epidermal growth factor receptor (EGFR) mutations and anaplastic lymphoma kinase (ALK) gene fusions. Drugs designed to target these drivers have dramatically improved outcomes for patients with the most common type of lung cancer, adenocarcinoma.

Yet despite these successes, driver mutations have been identified in only a minority of cases and patients with other types of lung cancer, such as squamous cell carcinoma (SCC) and small-cell lung cancer (SCLC), do not currently benefit from targeted therapies. As the leading cause of cancer- related mortality worldwide, better treatment options for patients with lung cancer have been and continue to be a pressing need.

Poster Child for Precision Medicine

Efforts to expand the pool of therapeutically actionable targets have turned to comprehensive molecular profiling studies. Technological advancements in our ability to detect these genetic variants in smaller samples and in minimally invasive techniques could help to bring the promise of precision medicine to a greater number of patients with lung cancer.Non—small cell (NSCLC) and SCLC histologies comprise the 2 major types of lung cancer, with the former making up 85% of cases. Among patients with NSCLC, nearly two-thirds have adenocarcinomas.

The prevalence of EGFR mutations in NSCLC is between 10% and 35%, depending on the patient population. Most commonly, the mutation occurs in exon 19 or 21 of the EGFR gene, activating the tyrosine kinase activity of the EGFR protein and promoting lung cancer growth and progression. The small-molecule EGFR inhibitors erlotinib (Tarceva) and gefitinib (Iressa) are accepted firstline therapies, demonstrating response rates up to 75% and improving progression-free survival (PFS), compared with standard chemotherapy regimens.

The more potent second-generation inhibitor afatinib (Gilotrif) is also approved as front-line therapy in EGFR-mutant metastatic NSCLC and demonstrated improved ORR and PFS compared with chemotherapy, but also improved overall survival (OS) in patients with exon 19 deletions.

The long-term benefit these drugs provide is hampered by the ability of cancer cells to evolve numerous different mechanisms of resistance. Approximately half of resistance cases involve a secondary mutation in the EGFR gene, T790M, the so-called gatekeeper mutation that restores ATP binding. A third generation of EGFR inhibitors with specificity for this EGFR mutant has been developed and osimertinib (Tagrisso) was very recently approved in this setting.

The discovery of gene fusions involving ALK, which result in constitutive activation of its tyrosine kinase activity, opened up another therapeutic avenue for patients with NSCLC. This abnormality is seen in 2% to 7% of patients and renders tumors sensitive to small-molecule inhibitors of the ALK protein. Crizotinib (Xalkori), although originally developed as a MET inhibitor, was subsequently found to have potent inhibitory activity against the ALK and ROS1 proteins and received regulatory approval following demonstration of improved response rates and PFS.

A similar narrative of acquired resistance has played out for ALK inhibitors, including an analogous gatekeeper mutation, L1196M, as the most common cause. The second-generation inhibitors ceritinib (Zykadia) and alectinib (Alecensa) have also been approved by the FDA, with alectinib showing a unique ability to benefit patients with central nervous system metastases.

The identification of gene fusions involving the ROS1 gene has further expanded the list of targetable oncogenic drivers in NSCLC, with the approved indications of crizotinib expanded to include patients with this rarer alteration in the past year.

Sequencing Studies Define Subtype Differences

Screening patients for these specific alterations and matching them to targeted therapy demonstrates the power of precision medicine in oncology. Yet, only a small proportion of patients with NSCLC are eligible for this kind of treatment; in the vast majority of cases, the molecular driver of the tumor remains unknown and 5-year survival rates are about 19%.Stand-alone studies have identified many potentially targetable mutations in NSCLC beyond EGFR and ALK, including other tyrosine kinase receptors such as MET and HER2 and downstream components of these signaling pathways such as BRAF, AKT, and PIK3CA, but none are currently actionable.

With the advent of next-generation sequencing, high-throughput molecular profiling studies have become a reality and enabled an even broader understanding of the potential drivers of a particular tumor type.

Adenocarcinoma

The lung cancer genome is characterized by a high frequency of somatic mutations of between 8 and 10 mutations per megabase, irrespective of the histologic subtype. That can make it challenging to pick out the driver mutations from the wealth of potential passenger mutations.The most commonly mutated gene in patients with adenocarcinomas is KRAS, encoding a GTPase enzyme that acts as a central node in numerous cellular signaling networks. Although a promising target, the RAS proteins have thus far eluded all efforts to design drugs that block their activity, but this remains an active area of ongoing research.

As expected, the EGFR gene is the next most significant driver gene in adenocarcinomas. A variety of other driver mutations, fusions, and amplifications have emerged from genome-wide sequencing studies, including those mentioned above.

Most recently, a large number of adenocarcinoma samples were profiled by The Cancer Genome Atlas (TCGA) Research Network. Analysis of tumor and matched normal samples from 230 previously untreated patients with lung adenocarcinoma revealed 18 significantly mutated genes.

In addition to the usual suspects, a high frequency of TP53 mutations (46%), as well as mutations in other tumor suppressor genes, such as NF1 (11%), KEAP1 (17%), STK11 (17%), and SMARCA4 (6%) were also noted.

The study also found that NF1, MET, HER2, and RIT1 mutations occurred in approximately 13% of patients collectively who otherwise did not display any driver mutations, suggesting that these could represent a novel set of targets in a unique subset of patients who cannot currently be treated with precision medicine.

In recent years, the MET gene has emerged as one of the most promising alternative actionable driver events in NSCLC. Amplification of the MET gene is found in about 2% to 4% of previously untreated patients with adenocarcinoma and has been shown to confer increased sensitivity to small-molecule inhibitors of MET.

MET amplification is also a common mechanism of resistance to EGFR inhibitors, allowing the cancer cell to bypass the need for EGFR by activating the MET signaling pathway to achieve the same ends. But development of these drugs in NSCLC has been plagued with problems.

Mutations in the MET gene have also been uncovered and a particular type of mutation is emerging as a promising new biomarker for MET inhibitor therapy. Known as exon 14 skipping mutations, they result in the juxtamembrane region, responsible for the proper degradation of the MET protein, being spliced out of the protein-coding region. Three different MET inhibitors have shown activity against tumors with MET exon 14 deletions, suggesting a potential route to approval for MET inhibitors in NSCLC.

Although the majority of lung cancers occur in those exposed to cigarette smoke, 10% to 20% of lung cancer cases in the United States occur in never-smokers and, as the rates of smoking decrease, these patients will begin to represent an increasing proportion of the tumor type.

Genome sequencing studies have revealed that the alterations underlying adenocarcinoma differ between smokers and non-smokers. The mutational burden in patients who have never smoked is typically much lower (0.8-1 mutations/ Mb) and they exhibit far fewer of the cytosine-adenine transversions that are a common feature of smoking-associated cancers.

The types of mutations are also quite distinct. Mutations in tumor suppressor genes, including TP53 and NF1, as well as mutations in KRAS, BRAF, and PIK3CA are more common in smokers, while EGFR and HER2 mutations and ALK, ROS1, and RET fusions were more frequently observed in never-smokers.

Squamous Cell Carcinoma

Finally, the number of neoepitopes, protein fragments that may be recognized by the immune system, was also significantly lower in neversmokers. This marker serves as an indication of the immunogenic potential of the mutational landscape of a tumor.SCCs are the second-largest group of NSCLCs and several studies had previously hinted that they have quite a different set of underlying genomic alterations from adenocarcinomas.

The TCGA performed the first comprehensive genome-wide study in 2012 and confirmed that this was indeed the case. EGFR mutations, ALK fusions, and other markers of adenocarcinomas were rare in SCC. Instead, SCCs were characterized by almost universal TP53 mutations, as well as common mutations in CDKN2A, PTEN, PIK3CA, KEAP1, MLL2, HLA-A, NFE2L2, NOTCH1, and RB1. Another common finding in SCCs is amplification of the gene encoding the fibroblast growth factor receptor (FGFR).

Interestingly, in terms of their genomic features, SCCs share more in common with head and neck squamous cell carcinomas and bladder cancers than with lung adenocarcinomas, which themselves are more similar to glioblastomas and colorectal cancers.

Small-Cell Lung Cancer

Although an effective targeted therapy has yet to be approved for the treatment of the SCC subtype, immunotherapy has emerged as a promising therapeutic strategy. A recent study directly comparing the genomic alterations in SCC and adenocarcinoma found that both had high levels of neoepitopes—47% in adenocarcinoma and 53% in SCC—reinforcing the potential for immunotherapy.SCLC is a particularly deadly form of lung cancer and has also thus far been resistant to targeted therapy. Genome sequencing studies have proved challenging because patients rarely undergo surgery, meaning that tumor specimens are lacking, but those that have been performed have revealed both commonalities and distinctions between SCLC and the 2 common types of NSCLC.

A key feature of SCLC is the nearly universal inactivation of the tumor suppressors TP53 and RB1, with mutation rates across several different studies of up to 90% and 65%, respectively. Thus, TP53 mutations are a common feature of all 3 major types of lung cancer, although they are much more common in SCLC and SCC than in adenocarcinoma.

Other likely driver mutations of SCLC include EP300, CREBBP, and various members of the Notch signaling pathway.

The SOX proteins have also been implicated in the development of SCLC, which display mutations, copy number alterations and amplification of various SOX gene family members. These proteins have diverse biological roles, but amplified SOX2 was highlighted in particular as a result of its role in stem cell biology.

Identifying Target Patients

The path to novel therapeutic options in SCLC is less clear, but immunotherapy is also demonstrating some efficacy in this tumor type, with nivolumab (Opdivo) and ipilimumab (Yervoy) both undergoing phase III clinical testing. The Notch 2/3 inhibitor OMP-59R5 is also being evaluated in a phase I/II clinical trial.Another significant challenge to the effective implementation of precision medicine in lung cancer is the need for optimal companion diagnostics to identify patients whose tumors carry actionable genomic alterations.

EGFR mutations can be detected by a number of different methods, each having advantages and disadvantages. The preferred method is the use of real-time polymerase chain reaction (RT-PCR) assays. In this category, the Therascreen EGFR RGQ PCR Kit is approved for use with afatinib and the cobas EGFR Mutation Kit with erlotinib, while the cobas EGFR Mutation Test v2 is designed to aid in the selection of patients with the EGFR T790M mutation who might benefit from osimertinib treatment.

Until recently, these tests were performed solely on tumor tissue samples. However, obtaining a sample of adequate quality and quantity can be challenging for several reasons, including the fact that relatively few patients undergo surgical resection and that both the purity and cellularity of a sample can be low for lung tumors since they are often infiltrated by other cell types.

The ability to gather tumor DNA from other sources, such as blood, serum, and plasma samples that may contain circulating tumor cells or cell-free DNA that can be used for genotyping is also being examined. So-called “liquid biopsies” offer several advantages; notably, they are minimally invasive and could be of particular benefit in patients who are very sick or where it is impossible to obtain adequate tumor tissue samples.

In June, the FDA approved the first blood-based companion diagnostic to detect EGFR mutations in patients with advanced NSCLC as an additional use for the cobas EGFR Mutation Test v2, and to determine eligibility for treatment with erlotinib.

The presence of ALK fusion genes is determined by detection of the protein product using either fluorescence in situ hybridization (FISH) or immunohistochemistry (IHC) techniques. Both types of tests give highly concordant results and both are approved by the FDA as companion diagnostics for ALK inhibitors but, although IHC is generally simpler to use and cheaper, FISH is still often regarded as the gold standard.

Immunotherapy Advances

Ultimately, the aim is to develop targeted next-generation sequencing panels that would allow the detection of multiple genomic alterations with a single test, but several challenges must be overcome before this type of testing becomes standard clinical practice.Although targeted therapy and chemotherapy remain the frontline standards for patients with advanced NSCLC depending on mutation status and tumor histology, the rapid development of checkpoint blockade immunotherapies may alter that paradigm.

In October, the PD-1 inhibitor pembrolizumab (Keytruda) became the first checkpoint blockade agent approved for the first-line treatment of patients with NSCLC whose tumors have ≥50% PD-L1 expression after demonstrating a 40% reduction in the risk of death for this population. The agent also is approved across all NSCLC histologies as second-line therapy for patients with ≥1% PD-L1 expression.

Nivolumab, which also targets PD-1, gained indications as a second-line treatment for patients with advanced squamous and nonsquamous NSCLC without a PD-L1 testing requirement. Additionally, the FDA has approved the PD-L1 inhibitor atezolizumab (Tecentriq) for patients with metastatic NSCLC who have progressed on prior therapies regardless of histology or PD-L1 expression status.

As the field develops, a robust debate on the merits of frontline targeted therapies versus immunotherapies is likely to unfold.

Key Research

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