Current Challenges in Molecular Testing in NSCLC

Lung cancer continues to be the leading cause of cancer-related mortality, resulting in ~1.4 million annual deaths worldwide and 160,000 deaths each year in the United States.¹However, in the past decade, some significant advances have been made in our understanding of the molecular mechanisms involved in lung cancer and, consequently, in the development of new therapeutic strategies.^1,2Indeed, while lung cancers were traditionally classified as either small cell lung cancers (SCLC) or non-small cell lung cancers (NSCLC), the selection of therapy is now guided by further subtyping in NSCLC using molecular testing; such testing allows for assessment of diagnosis and prognosis, as well as the eligibility of the patient for targeted therapies.²In particular, genetic alterations in the genes that encode two tyrosine kinases, the epidermal growth receptor (EGFR) and the anaplastic lymphoma kinase (ALK), are now known to play an integral role in lung cancer, providing an opportunity for targeted therapeutic interventions, which may offer substantive benefits over conventional chemotherapies.¹

Several practical challenges are associated with molecular testing in NSCLC, most notably, the collection and handling of the surgical pathology and/or cytopathology specimen, which may be the only material available for analysis.²It is also notable that modern, noninvasive biopsy procedures have unfortunately resulted in increasingly smaller samples for analysis. Although a lung resection will provide more than ample material for molecular analysis, only a small subset of patients will require a resection, so it cannot be assumed that a specimen will be available beyond the initial biopsy.²

Effective communication between the surgical pathologist and the treating physician is therefore essential to optimize care and maximize treatment choices (eg, establishing a priority for molecular and/or other types of testing if biopsy material is limited). Other important practical tasks to consider are the selection of an appropriate testing laboratory; Clinical Laboratory Improvement Amendment (CLIA) accreditation, for example, is especially important for diagnostic laboratories in the US.

A recently released document supports the use of molecular testing forEGFRmutations andALKfusions to guide patient selection for therapy with EGFR- or ALK-inhibitor drugs; these recommendations prioritize testing for these mutations over other molecular predictive tests and apply to all patients with advanced-stage adenocarcinoma, irrespective of clinical risk factors.¹Although only a subset of patients may have a molecular profile that is associated with response to an approved agent (eg,EGFR-activating mutation, orALKgene rearrangements), the detection of such alterations leads to a change in treatment.²Tyrosine kinase inhibitors (TKIs) such as gefitinib and erlotinib, which targetEGFR, have been an effective therapeutic interventions for NSCLC patients, although it is now known that not all patients benefit from this type of therapy. Accordingly, the identification of molecular markers of response has been an active area of research.^2,3The use of both negative and positive predictive biomarkers for response has been suggested as a means of maximizing the cost-effectiveness and therapeutic index of anti-EGFR therapy.³In this regard, negative biomarkers (ie, those that would exclude patients from anti-EGFR therapy) include negative immunohistochemical (IHC) staining for EGFR, KRAS mutation, and MET amplification, while positive predictive markers for response to therapy includeEGFRmutation and high copy gene number.³

An example of the impressive potential of molecular testing for NSCLC was recognized in 2007 with the identification of a 5-gene signature that could predict increased recurrence risk and reduced overall survival in patients with surgically resected NSCLC.⁴The gene signature was strongly associated with survival, with a sensitivity of 98%, a specificity of 93%, and an overall accuracy of 96%.⁴Although the 5-gene signature must also be validated in large-scale, multicenter, prospective studies, it ultimately may be useful to identify patients with NSCLC at risk for an aggressive clinical course, who might therefore benefit from adjuvant chemotherapy; conversely, those patients in the low-risk group who could be spared unnecessary treatment.^4,5The different genes identified in the signature include signal transducer and activator of transcription 1 (STAT1); monocyte to macrophage differentiation-associated protein (MMD); dual specificity phosphatase 6 (DUSP6); v-erb-b2 avian erythroblastic leukemia viral oncogene homolog 3 (ERBB3); and lymphocyte-specific protein tyrosine kinase (LCK). That range of genes might also serve as useful targets to develop additional drug therapies.^4,5Thus, althoughEGFRmutations,ALKfusions, and, frequently,KRASmutations (since the latter serves as a negative predictor of response to targeted therapy) are currently the most important recommended targets, other mutations that, based on clinical data, might one day be added to the list of mutations to evaluate include targets such asRos1,HER2,BRAF, andPIK3CA.²

A number of challenges associated with the development of guidelines for molecular testing in NSCLC have also been identified, and these include the sheer diversity of molecular alterations that need to be assessed, an increasing number of clinically relevant cancer-associated genes, and the limited literature and testing experience associated with newly developed targeted therapies and with their accompanying diagnostic tests.¹Nonetheless, there will continue to be a need for guidance statements such as those from the College of American Pathologists, the International Association for the Study of Lung Cancer, and the Association for Molecular Pathology to establish literature-based consensus for best practice diagnostic procedures in NSCLC.

George R. Simon, MD

Commenting on this issue, George R. Simon, MD, professor of Medicine and Oncology and section chief of Translational Research at the Department of Thoracic/Head & Neck Medical Oncology at MD Anderson Cancer Center in Houston, Texas, noted that, for NSCLC patients: “I think it is very important to do a detailed molecular profile because it has direct therapeutic implications.” For patients with actionable targets, Simon suggested that they should be treated specifically with the targeted agent; conversely, Simon noted, “If there are no directly targetable agents immediately available, then the patient should be referred for an appropriate clinical trial.”

Simon also pointed out some of the important challenges associated with molecular testing in NSCLC, specifically, the challenges associated with obtaining sufficient tissue for molecular analysis, and the time needed to both schedule a biopsy (~ 1 week) and then waiting for the results, which could be on the order of 2 to 3 weeks. He said that occasionally up to a month or more could elapse between scheduling the biopsy and receiving the actionable data.

Other patient-specific issues can lead to further delays; patients on blood thinners, for example, will need to be taken off these medications for several days before a biopsy can even be done. In addition, the molecular analysis results may come back with no actionable data, leading to a considerable delay in starting appropriate therapy for patients.

Regarding the number of molecular targets available to screen, Simon said that while private testing panels are available from some companies for up to 250 genes at a considerable cost, most companies and institutions, including his own, will screen for a panel of 50 or so genes, but only charge forEGFR,KRAS, andALK, despite running a panel of many more genes. Simon citedRos1, RET,KRAS, and, more recently, NTRK, as targets that may be added to the list of molecular targets in NSCLC.

Vali A. Papadimitrakopulou, MD, professor in the Department of Thoracic/Head and Neck Medical Oncology at the MD Anderson Cancer Center, explained that there are many additional and potentially important genomic alterations in NSCLC, but that validation of their role as actual drivers of the oncogenic and/or metastatic process is lagging. He also noted that most of the genomic alterations that have been used to define potential targets for therapy, and ultimately drug approval, have been largely confined to the setting of lung adenocarcinoma, while other histolopathologies, including squamous cell lung cancer, SCLC, and mesothelioma, have been largely under-represented. As such, one area of high interest is the recognition of the role of genomic alterations in squamous cell lung cancer, and the potential to target these alterations therapeutically in large-scale clinical trials that will use next-generation sequencing (NGS) platforms as the screening tool. In this regard, he referred to a national effort, the Phase II/III Biomarker-Driven Master Protocol for Second Line Therapy of Squamous Cell Lung Cancer.6 Patients in this study will be genotyped using NGS and IHC methods, and based on the resulting profile, placed into one of multiple arms of a clinical trial. In the context of squamous cell carcinoma, which constitutes about 25% of all lung cancers, some two dozen compounds that have shown measurable activity against some 16 different molecular targets will be open to investigation.⁶The trial differs from other biomarker-driven trials in that prespecified criteria have been defined, with the goal of facilitating US Food and Drug Administration (FDA) approval of qualifying compounds. In addition, the trial has a modular design, which, based on evolving results, allows for one arm to be dropped in favor of another.⁶Although now basically a ‘test drive’ with squamous cell carcinoma, this type of initiative, if successful, could be applied more broadly to NSCLC, as well as to many other cancer types. Commenting on the benefit of such initiatives, Papadimitrakopulou said: “We hypothesize that this Master Protocol mechanism will yield definable and measurable efficiencies in terms of improving genomic screening of cancer patients for clinical trial entry, and improved time lines for drug-biomarker testing, allowing for inclusion of the maximum numbers of otherwise-eligible patients in comparison with currently employed ‘single screen-single trial’ approaches.”

References

1. Lindeman NI, Cagle PT, Beasley MB, et al. Molecular testing guideline for selection of lung cancer patients for EGFR and ALK tyrosine kinase inhibitors: guideline from the College of American Pathologists, International Association for the Study of Lung Cancer, and Association for Molecular Pathology.J Thorac Oncol. 2013;8(7):823-859.
2. Aisner DL, Marshall CB. Molecular pathology of non-small cell lung cancer: a practical guide.Am J Clin Pathol. 2012;138(3):332-346.
3. John T, Liu G, Tsao MS. Overview of molecular testing in non-small-cell lung cancer: mutational analysis, gene copy number, protein expression and other biomarkers of EGFR for the prediction of response to tyrosine kinase inhibitors.Oncogene. 2009;Aug;28(suppl 1):S14-S23.
4. Chen HY, Yu SL, Chen CH, et al. A five-gene signature and clinical outcome in non-small-cell lung cancer.N Engl J Med. 2007;356(1):11-20.
5. Swisher SG, Kuerer HM. Commentary: five-gene signature for survival in non—small-cell lung cancer patients. http://www.jwatch.org/oh200701290000001/2007/01/29/five-gene-signature-survival-non-small-cell-lung. Accessed February 3, 2014
6. The ASCO Post: 'Master Protocol' could revolutionalize trials in lung cancer, and eventually other cancers. http://www.ascopost.com/issues/november-1,-2013/master-protocol-could-revolutionalize-trials-in-lung-cancer,-and-eventually-other-cancers.aspx. Accessed February 10, 2014.