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The Utilization of Cytologic Fine-Needle Aspirates of Lung Cancer for Molecular Diagnostic Testing.

Roh MH - J Pathol Transl Med (2015)

Bottom Line: In this era of precision medicine, our understanding and knowledge of the molecular landscape associated with lung cancer pathogenesis continues to evolve.During the management of these patients, minimally invasive procedures to obtain samples for tissue diagnoses are desirable.Thus, cytologic fine-needle aspirates must be utilized and triaged judiciously to achieve both objectives.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, University of Michigan Health System, Ann Arbor, MI, USA.

ABSTRACT
In this era of precision medicine, our understanding and knowledge of the molecular landscape associated with lung cancer pathogenesis continues to evolve. This information is being increasingly exploited to treat advanced stage lung cancer patients with tailored, targeted therapy. During the management of these patients, minimally invasive procedures to obtain samples for tissue diagnoses are desirable. Cytologic fine-needle aspirates are often utilized for this purpose and are important not only for rendering diagnoses to subtype patients' lung cancers, but also for ascertaining molecular diagnostic information for treatment purposes. Thus, cytologic fine-needle aspirates must be utilized and triaged judiciously to achieve both objectives. In this review, strategies in utilizing fine-needle aspirates will be discussed in the context of our current understanding of the clinically actionable molecular aberrations underlying non-small cell lung cancer and the molecular assays applied to these samples in order to obtain treatment-relevant molecular diagnostic information.

No MeSH data available.


Related in: MedlinePlus

Examples of epidermal growth factor receptor (EGFR) mutations detected by the polymerase chain reaction based fragment analysis assay utilized at our institution. Our assay is a multiplex assay designed to detect the two most common mutations in EGFR: the L858R substitution (case 1) and small deletions within exon 19 (case 2).
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f2-jptm-49-4-300: Examples of epidermal growth factor receptor (EGFR) mutations detected by the polymerase chain reaction based fragment analysis assay utilized at our institution. Our assay is a multiplex assay designed to detect the two most common mutations in EGFR: the L858R substitution (case 1) and small deletions within exon 19 (case 2).

Mentions: EGFR mutation analysis is commonly performed via polymerase chain reaction (PCR) and sequencing-based approaches; advances in the development of testing modalities have afforded a multitude of methodologies [1,17,18]. Sanger sequencing is considered the gold standard as this involves direct DNA sequence acquisition and can provide information regarding the presence of all potential mutations including common, known mutations and novel mutations. Nonetheless, this test requires a relatively higher enrichment of tumor cell DNA content in the sample. The typical analytic sensitivity for Sanger sequencing is 15%–20% mutant allele, which equates to 30%–40% tumor cells assuming that the genetic mutation is a heterozygous event without amplification [17]. This can be problematic in both small biopsy and cytology specimens, especially cell blocks, in which the tumor cell population can be diluted by background benign cellular elements such as inflammatory cells, bronchial epithelial cells, and/or stromal mesenchymal cells. Especially in this setting, a negative mutation result can be either due to the true absence of the mutation in the tumor cells or insufficient percent tumor cellularity that falls below the analytic sensitivity threshold thereby resulting in the failure to detect the mutation even despite the presence of the mutation [4,9,17,19]. Therefore, often times, there is more reliance on tumor cell enrichment by either macrodissection or microdissection to obtain a reliable result [7,9,17,20]. Sanger sequencing is also relatively more labor intensive and time consuming than targeted methods and can lead to longer turnaround times [18]. In contrast to the general Sanger sequencing approach, targeted mutation detection methods such as PCR-restriction fragment length polymorphism, real-time PCR, pyrosequencing, high resolution melting analysis (HRMA), and PCR fragment analysis can be utilized [17]. The advantages of these approaches include their improved analytic sensitivity and less time-consuming nature leading to reduced turnaround times. At our institution, we utilize a multiplex PCR fragment analysis assay for EGFR mutation testing; this allows for the simultaneous assessment of the two most commonly observed EGFR mutations (Fig. 2). The analytic sensitivity of this method is better than that of Sanger sequencing; a minimum of only 10% tumor cells is required. In the past decade, myriad studies have been reported demonstrating that a variety of cytologic samples and cytopreparatory platforms can be effectively utilized for EGFR mutational analysis. These have been reviewed elsewhere [18,21] but salient examples will be discussed below.


The Utilization of Cytologic Fine-Needle Aspirates of Lung Cancer for Molecular Diagnostic Testing.

Roh MH - J Pathol Transl Med (2015)

Examples of epidermal growth factor receptor (EGFR) mutations detected by the polymerase chain reaction based fragment analysis assay utilized at our institution. Our assay is a multiplex assay designed to detect the two most common mutations in EGFR: the L858R substitution (case 1) and small deletions within exon 19 (case 2).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4508567&req=5

f2-jptm-49-4-300: Examples of epidermal growth factor receptor (EGFR) mutations detected by the polymerase chain reaction based fragment analysis assay utilized at our institution. Our assay is a multiplex assay designed to detect the two most common mutations in EGFR: the L858R substitution (case 1) and small deletions within exon 19 (case 2).
Mentions: EGFR mutation analysis is commonly performed via polymerase chain reaction (PCR) and sequencing-based approaches; advances in the development of testing modalities have afforded a multitude of methodologies [1,17,18]. Sanger sequencing is considered the gold standard as this involves direct DNA sequence acquisition and can provide information regarding the presence of all potential mutations including common, known mutations and novel mutations. Nonetheless, this test requires a relatively higher enrichment of tumor cell DNA content in the sample. The typical analytic sensitivity for Sanger sequencing is 15%–20% mutant allele, which equates to 30%–40% tumor cells assuming that the genetic mutation is a heterozygous event without amplification [17]. This can be problematic in both small biopsy and cytology specimens, especially cell blocks, in which the tumor cell population can be diluted by background benign cellular elements such as inflammatory cells, bronchial epithelial cells, and/or stromal mesenchymal cells. Especially in this setting, a negative mutation result can be either due to the true absence of the mutation in the tumor cells or insufficient percent tumor cellularity that falls below the analytic sensitivity threshold thereby resulting in the failure to detect the mutation even despite the presence of the mutation [4,9,17,19]. Therefore, often times, there is more reliance on tumor cell enrichment by either macrodissection or microdissection to obtain a reliable result [7,9,17,20]. Sanger sequencing is also relatively more labor intensive and time consuming than targeted methods and can lead to longer turnaround times [18]. In contrast to the general Sanger sequencing approach, targeted mutation detection methods such as PCR-restriction fragment length polymorphism, real-time PCR, pyrosequencing, high resolution melting analysis (HRMA), and PCR fragment analysis can be utilized [17]. The advantages of these approaches include their improved analytic sensitivity and less time-consuming nature leading to reduced turnaround times. At our institution, we utilize a multiplex PCR fragment analysis assay for EGFR mutation testing; this allows for the simultaneous assessment of the two most commonly observed EGFR mutations (Fig. 2). The analytic sensitivity of this method is better than that of Sanger sequencing; a minimum of only 10% tumor cells is required. In the past decade, myriad studies have been reported demonstrating that a variety of cytologic samples and cytopreparatory platforms can be effectively utilized for EGFR mutational analysis. These have been reviewed elsewhere [18,21] but salient examples will be discussed below.

Bottom Line: In this era of precision medicine, our understanding and knowledge of the molecular landscape associated with lung cancer pathogenesis continues to evolve.During the management of these patients, minimally invasive procedures to obtain samples for tissue diagnoses are desirable.Thus, cytologic fine-needle aspirates must be utilized and triaged judiciously to achieve both objectives.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, University of Michigan Health System, Ann Arbor, MI, USA.

ABSTRACT
In this era of precision medicine, our understanding and knowledge of the molecular landscape associated with lung cancer pathogenesis continues to evolve. This information is being increasingly exploited to treat advanced stage lung cancer patients with tailored, targeted therapy. During the management of these patients, minimally invasive procedures to obtain samples for tissue diagnoses are desirable. Cytologic fine-needle aspirates are often utilized for this purpose and are important not only for rendering diagnoses to subtype patients' lung cancers, but also for ascertaining molecular diagnostic information for treatment purposes. Thus, cytologic fine-needle aspirates must be utilized and triaged judiciously to achieve both objectives. In this review, strategies in utilizing fine-needle aspirates will be discussed in the context of our current understanding of the clinically actionable molecular aberrations underlying non-small cell lung cancer and the molecular assays applied to these samples in order to obtain treatment-relevant molecular diagnostic information.

No MeSH data available.


Related in: MedlinePlus