The Daily Sign-Out

Mutations in Lung Adenosquamous Carcinomas

Journal of Thoracic Oncology just issued a published ahead-of-print article that examined mutations of most frequent driver mutations seen in pulmonary adenocarcinomas in a series of 76 resected adenosquamous carcinomas of the lung.

This is an important study to note because it supports the recommendation by IASLC/ERS/ATS to send these tumors for molecular testing.  Another interesting finding for pathologists is the finding of excess ALK mutations associated with those tumors with a solid pattern histologic subtype in the adenocarcinoma component.

 

Analysis of Major Known Driver Mutations and Prognosis in Resected Adenosquamous Lung Carcinomas

Wang, Rui MD; Pan, Yunjian MD; Li, Chenguang MD,PhD; Zhang, Huibiao MD; Garfield, David MD; Li, Yuan MD; Ye, Ting MD; Hu, Haichuan MD; Luo, Xiaoyang MD; Li, Hang MD; Zhang, Yang MD; Zhang, Jie MD; Zhou, Xiaoyan MD; Shen, Lei MD; Pao, William MD,PhD; Sun, Yihua MD; Chen, Haiquan MD

Published Ahead-of-Print

 

Abstract

 

Introduction: Genotyping for driver mutations is now routinely used to guide clinical care of patients with lung cancer. Adenosquamous lung carcinoma (AdSqLC) is a subtype of cancer that contains both adenocarcinoma and squamous cell carcinoma. However, the incidence, clinicopathologic characteristics, and prognostic implications of major driver mutations in AdSqLCs are not well established.

Methods: Seventy-six resected AdSqLCs and 646 lung adenocarcinomas were screened for known genetic alterations involving EGFR, ERBB2, KRAS, BRAF, PIK3CA, AKT1, RET, and ALK. Tumors showing acinar, lepidic, micropapillary, or papillary growth in glandular component were classified as classical AdSqLC.

Results: Of the 76 AdSqLCs, 43 (56.6%) harbored known mutant kinases, including 24 (31.6%) with EGFR mutations, eight (10.5%) with KRAS mutations, two (2.6%) with AKT1 (2.6%) mutations, one (1.3%) with ERBB2 insertion mutation, one (1.3%) with PIK3CA mutation, four (5.3%) with ALK fusions, and three (4%) with KIF5B-RET fusions. No mutation was found in BRAF. The mutational profiles and clinicopathologic characteristics of classical AdSqLC were strikingly similar to that of poorly differentiated adenocarcinoma. However, AdSqLCs with solid growth pattern in glandular component had high frequency of ALK or RET fusions and low EGFR mutation rate.

Conclusions: To our knowledge, this is the first comprehensive study investigating major oncogenic driver mutations in a large cohort of AdSqLC patients in a Chinese population. The findings suggest that it will be clinically valuable to investigate the growth pattern of glandular component in AdSqLCs.

ETS2 mediates tumor suppressor function and MET inhibition in lung adenocarcinoma

There was a very intriguing paper published online by Clinical Cancer Research on May 9 last week that should be of interest for those involved in lung cancer research.  The authors are Mohamed Kabbout, Melinda Garcia, Junya Fujimoto, et al. from M.D. Ancerson Cancer Center in Houston and the title of the article is "ETS2 mediated tumor suppressive function and MET oncogene inhibition in
human non-small cell lung cancer." 

In brief, the authors find that there is significant downregulation of the ETS2 transcription factor in lung adenocarcinomas compared to normal lung tissues.  ETS2 expression is linked to various canonical cancer-associated pathways in lung cancer cells, in particular the HGF pathway.  Moreover, knockdown of ETS2 is associated with up-regulation of the ZEB1, SNAI2, RAB3B, SERPINB5 and SPDEF genes
with concomitant with elevated cell migration and invasion.  Last, low ETS2 protein is predictive of shorter time-to-recurrence in NSCLC and lung adenocarcinoma.  These data suggest that ETS2 functions as a tumor-suppressor in lung cancer cells by, at least in part, regulation of the migration- and invasion-promoting transcriptome.

More details. . .

Gene expression profiling showed significant decreased expression of ETS2 transcription factor transcripts in lung adenocarcinoma (ADC) compared to normal controls and quantitative RT-PCR confirmed decreased ETS2 expression.  Subset analysis should significantly decreased ETS2 expression in those ADCs from smokers compared to those from never-smokers.

They analyzed ETS2 expression by IHC in a microarray set of 201 NSCLC (135 ADC, 66 squamous cell carcinoma-SQC) tumors.  Low ETS2 expression was associated with shorter time-to-recurrence in stage I and all stages of patients.  Multivariate Cox proportional hazards analysis showed that, after adjusting for stage, low ETS2 expression was an independent predictor of shorter time-to-recurrence in NSCLC and ADC.

H441 human lung cancer cell lines transfected with small interfering RNA (siRNA) specific to ETS2 showed significantly reduced ETS2 transcript compared to controls.  ETS2 knockdown significantly increased cell growth, migration, and invasion.  Conversely, ETS2 overexpression in H1299 lung cancer cells showed significantly decreased cell migration and invasion.  Of interest, the magnitude of these changes following knockdown or overexpression of ETS2 was greater in measures of migration and invasion compared to cell growth--suggesting a tumor suppressor function for ETS2 at least in vitro.

Gene expression profiling of H441 lung cancer cell lines transfected with scrambled siRNA (control) versus ETS2-specific siRNA showed significant differential expression in 1,816 transcripts.  Functional analysis by Ingenuity Pathways Analysis (IPA) showed ETS2 knockdown modulated key oncogenic pathways, including HGF, integrin, tissue factor, semaphorin, and MAPK signaling.  Notably, HGF was the top modulated canonical pathway identified following knowdown, and further, an HGF-mediated gene-interaction network was among the among the top significant gene networks following topological arrangement of the genes found to be differentially expressed by IPA.

Western blot analysis and ELISA both showed increased phosphorylation MET following ETS2 knockdown.  Importantly, co-transfection of H441 lung cancer cells with MET-specific siRNA abrogated cell migration, invasion, and, to a lesser effect, cell growth mediated by knockdown of ETS2 expression alone.  Furthermore, co-transfection of cells with MET-specific siRNA significantly attenuated the induction of SNAI2, RAB3B, SPDEF and SERPINB5 mediated by knockdown of ETS2 alone as well as reduced expression of ZEB1. These data demonstrate that ETS2 suppresses key features of the lung cell malignant phenotype, at least in part, through inhibition of the MET oncogene. 

The authors also find that ETS2 suppresses EGF-mediated signaling in lung cancer cells.  HGF treatment increased ETS2 protein expression and phosphorylation of MET and ERK1/2 MAPK in H441 cells.  Conversely, expression of ETS2, phospho-MET, and phospho-ERK1/2 was attenuated by co-treatment of H441 cells with the MET TKI PHA-665752.  HGF treatment also significantly increased the expression of SNAI1, RAB3B, SPDEF, ZEB1, and SERPINB5 genes.  Moreover, the expression of these genes was significantly higher in HGF-treated cells transfected with ETS2-specific siRNA compared to similarly treated control cells.  Conversely, overexpression of ETS2 in H1299 cells decreased HGF-induced phosphorylated MET levels and decreased HGF-induced cell migration in the wound healing assay. 

Finally, NSCLC (p=0.01) or ADC (p=0.02) patients with relatively low ETS2 protein expression and higher phospho-MET membrane immunoreactivity showed shortest tiime-to-recurrence compared to other patient groups, specifically those with higher ETS2 and higher phospho-MET
expression.

NKX2-1/TTF-1 drives pulmonary-specific differentiation in lung adenocarcinoma

Snyder et al. recently published an intriguing report in Molecular Cell (21 March 2013) (featured free article) in which they convincingly demonstrate how tumors in which Nkx2-1 was deleted show striking different morphology from those in which Nkx2-1 is expressed.

Nkx2-1/TTF-1 is a highly conserved homeodomain-containing transcription factor that is expressed at the onset of lung and thyroid development and has been shown from murine knockout experiments to be required for branching morphogenesis in very early lung development. Nkx2-1 is expressed in about 80% of human lung adenocarcinomas (ADCs). Recent studies have shown that Nkx2-1-negative ADCs have a worse prognosis and have a higher histologic grade (more poorly differentiated) compared to Nkx2-1-positive tumors. These data suggest that Nkx2-1 may coordinate a lineage-specific differentiation program on lung adenocarcinomas that restrains their malignant potential. Further data have shown that the Nkx2-1 gene is amplified in 10%–15% of human lung ADCs, suggesting that it can also act as a lineage-survival oncogene in a subset of tumors. The authors' previous work using a mouse model for lung ADC showed that Nkx2-1 restrained the ability of Kras-driven lung tumors to evolve to a poorly differentiated, Hmga2-positive state. 

The authors cleverly designed a mouse model in which Cre recombinase can activate a conditional allele of Kras ^G12D and delete both alleles of Nkx2-1 from the lungs of mice.  Simultaneous activation of Kras ^G12D and deletion of Nkx2-1 yielded peripheral lung ADCs that showed a distinct glandular growth pattern of malignant cells with abundant MUC5A-positive mucin resembling human lung mucinous ADC. In contrast, control Nkx2-1-positive mice developed tumors showing a predominantly papillary non-mucinous pattern. They also showed in a different model that in established tumors, Nkx2-1 deletion induced a glandular pattern and mucin production similar to de novo tumors and promoted long-term tumor growth. In addition, this study demonstrated that Nkx2-1 deletion induced diffuse alveolar epithelial hyperplasia in the alveolar space.  The hyperplastic cells lacked expression of canonical Nkx2-1 targets and exhibited increasing mucinous accumulation and morphology over time.

The authors compared Kras-mutant/Nkx2-1-deleted tumors and Nkx2-1-positive tumors by mRNA expression profiling and found 1,828 differentially expressed genes, including 1,137 upregulated and 691 downregulated genes. MetaCore analysis for disease-specific gene collections and networks identified genes associated with gastric diseases that were enriched in  ‘‘stomach neoplasms’’ and ‘‘stomach diseases’’ as categories that were enriched in Kras-mutant/Nkx2-1-deleted tumors.  Finally, they determined whether human mucinous lung adenocarcinomas also exhibit evidence of gastric differentiationby evaluating the expression of two stomach-restricted proteins (GKN1 and CTSE) in 37 human lung adenocarcinomas. In 11 NKX2-1-negative mucinous human lung adenocarcinomas (n = 11), they found that GKN1 was expressed in 6 out of 11 cases and that CTSE was strongly and diffusely expressed in all 11 tumors. In contrast, NKX2-1-positive lung adenocarcinomas (n = 26) were entirely negative for GKN1. Most NKX2-1-positive lung tumors were either negative (n = 14) or only focally positive (n = 10) for CTSE. 

The DL

There is much more to unpack from this article than I can concisely summarize in this post. The fascinating point for me is that lung alveolar epithelial cells have a latent gastric differentiation program that becomes activated by default when Nkx2-1/TTF-1 is deleted and this is most dramatically evinced in pulmonary mucinous adenocarcinomas. These particular tumors are nettlesome diagnostically because of the question raised of whether the tumor is primary or metastatic.

Wonderful paper and lots to think about!

 

Video: Dr. Jack West discusses changing views on molecular testing in NSCLC

The Lungevity blog features a video of Dr. Jack West from Swedish Cancer Institute in Seattle discussing his evolving views regarding molecular testing for actionable driver mutations in non-small-cell lung cancer (NSCLC). Dr. West is a thought-leader in lung oncology and I found this a tidy and brief summary of current issues and reasonable argument for expansion of such testing.

This is important for pathologists as we are often tasked with triaging testing as the time of diagnosis, managing results reporting, and driving testing. My own views as a pathologist with a concentration on lung cancer have changed over the last two years based on clinical practice, research meetings, recent publications, and listening to clinicians' views. Initially, it seemed to me that reflex testing of all advanced stage (stage 2B and up) lung adenocarcinomas and excluding tumors with squamous histology; since we generally are unaware of the smoking history, I ignored smoking history as a criterion for testing. I okayed this with the oncologists and off we went. Dr. West succinctly contrasts the two views concerning molecular testing in NSCLC: essentially testing all patients with advanced stage NSCLC versus testing focused on "enriched" populations, e.g. adenocarcinoma histology, never/light smokers.

My own views began to change as a result of one memorable lung cancer research meeting at Rush we had when one of the thoracic surgeons related how a patient grilled him on why he received an expensive bill for all this molecular testing without any discussion with the surgeon or oncologist and, moreover, when he had already expressed his preference to not receive any further treatment following biopsy and mediastinoscopy. These aren't cheap tests and someone (not the pathologist) must explain these results to the patient. I also reviewed our experience with molecular testing and found several instances where testing was sent on early-stage patients and squamous histology tumors, which I didn't think was appropriate. Based on another round of discussions with the oncologists, I decided that we should consult with the oncologist before ordering molecular testing.

But as Dr. West says, "Some patient's cancer don't read the books." He relates several recent experiences that have caused him to revise his views: squamous tumors that show driver mutations, including one tumor with both EGFR activating mutation and ALK rearrangement. He advocates broader, more expansive molecular testing as well as consideration of testing recurrences and re-biopsy of clinically stable lesions.

The bottom line is that this is still an evolving area--we must be aware of new data coming out and be prepared to modify our views and practices.

Lack of NKX2-1 (TTF-1) expression in lung adenocarcinoma and EGFR mutations

My last post discussed one of the two papers I thought were notably in the October 2012 Journal of Thoracic Oncology. Here's my take on the other paper by Vincentin et al (J Thorac Oncol 2012;7:1522-1527). NK2 homeobox 1 is the "proper" name that we pathologists more affectionately call "TTF-1." NKX2-1 is normally expressed in type 2 pneumocytes and non-ciliated bronchiolar epithelial cells and has been shown to play an active role in sustaining lung adenocarcinoma. Since NKX2-1 is found in about 95% of lung adenocarcinomas with EGFR mutations, the authors studied whether NKX2-1 IHC staining is a useful provisional marker for guiding EGFR-TKI therapy in advanced stage lung adenocarcinoma while EGFR mutation analysis is being completed.

This is a large study including 810 consecutive NSCLC tumor specimens referred for routine diagnostic EGFR mutation testing. While one may debate the ultimate practicality of this study with respect to it's purpose, what recommends it to me is that the study population more closely mirrors what a general pathologist encounters in routine practice. The study includes biopsies (N=594), resections (N=162), and cytology specimens (N=54) and the overall incidence of EGFR mutations was 14%. It is important to note that the authors used the TTF-1 8G7G3/1 antibody clone for IHC testing. There are substantial differences between the different antibody clones for TTF-1 that have been recently noted in the literature and it is important to take note of this detail when analyzing articles that test NKX2-1 or determining the best antibody to use in your lab.

Results and Authors' Conclusions

NKX2-1 was evaluable in 98% of cases and was positive in 68%, negative in 32%. NKX2-1 positive immunostaining showed a strong correlation with EGFR mutations: in the EGFR-mutated cases, NKX2-1 was positive in 92% and negative in 8%. The probability of absent of EGFR mutation in all adenocarcinomas is 80%; in NKX2-1-negative adenocarcinomas that probability rises to 96%. The authors found a negative predictive value of negative NKX2-1 IHC for EGFR mutation to be >95%.

The authors conclude that if treatment is clinically urgent, the NPV of NKX2-1 expression is appropriate to use as a surrogate marker to choose conventional chemotherapy as starting treatment pending EGFR mutation analysis. The PPV of NKX2-1 is not sufficient to warrent adding an EGFR-TKI. Also, the result of NKX2-1 is not specific enough to exclude a specimen from EGFR-mutation analysis.

My take-aways

While the clinical situation in which NKX2-1 testing would influence an urgent treatment decision may uncommon, I think it is important to note this paper because when it does occur, the pathologist can play a vital role in assisting the clinician determine the management for such a patient. If one is following the diagnostic algorithm from the 2011 IASLC guidelines, one would not routinely test either histologically diagnostic adenocarcinomas or squamous cell carcinomas for NKX2-1 (TTF-1). Another point is that the SPT24 clone for TTF-1 is a more sensitive antibody for TTF-1/NKX2-1 (a point that the authors make as well) and, therefore, one would expect the percentage of NKX2-1-negative cases to be fewer and the NPV to be higher. Finally, the authors used direct sequencing of EGFR exons 19-21, followed by HRM screening, and PCR-sequencing for identifying EGFR mutations; another methodology could influence the denominator of EGFR-mutated cases and, thus, the predictive value of NKX2-1 IHC testing.

Loss of PTEN as a poor prognostic marker for lung adenocarcinoma

The October 2012 Journal of Thoracic Oncology presents two papers of interest to pathologists.

The first study is a paper by Yanagawa et al. that correlates PTEN expression with gene copy number, P53 expression, EGFR and KRAS mutations in surgically resected NSCLCs and examines the relationship between PTEN status, clinicopathologic characteristics and survival.

Recall that PTEN functions as a lipid phosphatase that negatively regulates the PI3K/AKT anti-apoptotic and proliferation pathway, which is a key downstream component of the EGFR pathway. Loss of PTEN is most commonly due to promoter hypermethylation, while homozygous deletion and nonsense mutations with LOH seem to be uncommon.

PTEN expression was examined by immunohistochemistry in a tissue microarray constructed from 152 patients with a median follow-up time of 2.3 years. The study consisted of 62% adenocarcinomas, 29% squamous cell carcinomas and 9% of tumors called “other.” Since this is a study of surgically resected patients, there is obviously an overrepresentation of stage one tumors compared to all patients typically diagnosed with lung cancer. Smokers comprised 79% of the study population. EGFR mutations were found in 15% and KRAS mutations and 22%. The authors defined loss of PTEN as complete absence of cytoplasmic staining in tumor cells.

Loss of PTEN was found in 63 out of 152 cases (41.4%). No significant differences were found between loss of PTEN and smoking, EGFR mutations, KRAS mutations, or p53 expression.

Patients with loss of PTEN expression had significantly worse disease-free survival compared with patients with retained PTEN expression (HR 1.76, log-rank p=0.038) but the prognostic effect was related directly to tumor histology: loss of PTEN was significantly associated with shorter DFS in adenocarcinoma only (HR 2.72, log-rank p=0.002). This association was also significant in multivariate analysis controlling for sex, age, histology and stage.

Recent data suggest that loss of PTEN may contribute to EGFR TKI resistance in NSCLC. PIK3CA mutations only account for 2% to 4% of NSCLC but loss of PTEN would result in constitutive activation of the PI3K pathway. This study suggests that lung adenocarcinomas selected for PTEN status may be susceptible to PI3KCA inhibitors that are currently under investigation.

My next post will discuss the other interesting paper in JTO this month.

The New IASLC/ATS/ERS Lung Adenocarcinoma Classification

FYI-Excellent content for CME 
 
In 2011, the Journal for Thoracic Oncology published the "International Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society International Multidisciplinary Classification of Lung Adenocarcinoma," issued by a multidisciplinary team of lung cancer specialists.
 
To better understand the general implications of this new classification, and the issues affecting specific specialties, please join us in this series of webinars.


IASLC Experts Discuss Events
That Are Changing Clinical Practice

 
Module 1 - Introducing The New IASLC/ATS/ERS Lung Adenocarcinoma Classification
William D. Travis, MD
Memorial Sloan Kettering Cancer Center - NY, NY

Module 2 - Implications of the New IASLC/ATS/ERS Classification
Fred R. Hirsch, MD, PhD
University of Colorado Cancer Center - Aurora, CO
Paul Van Schil, MD, PhD
University Hospital of Antwerp - Edegem (Antwerp), Belgium
 
Module 3 - Highlights for the Pathologist
Ming S. Tsao, MD, FRCPC

Princess Margaret Hospital - Toronto, Ontario, Canada
 
Module 4 - Highlights for the Radiologist
Kavita Garg, MD
University of Colorado Cancer Center - Aurora, CO
 
Module 5 - Case Studies for the Practicing Clinician
Gregory Riely, MD, PhD
Memorial Sloan Kettering Cancer Center - NY, NY













This activity has been planned and implemented in accordance with the Essential Areas and policies of the Accreditation Council for Continuing Medical Education through the joint sponsorship of the International Association for the Study of Lung Cancer (IASLC) and the University of Colorado School of Medicine. The University of Colorado School of Medicine is accredited by the ACCME to provide continuing medical education for physicians.

 

                       This activity is supported in part, by an educational grant from Eli Lilly and Company










 



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Implications of clonality in multifocal non small cell lung cancer

About 5% to 8% of NSCLC patients present with 2 or more anatomically separate lung tumors at diagnosis. The distinction between synchronous primary tumors and intrapulmonary metastases has important implications for staging and, thus, treatment strategies.

Arne Warth and colleagues from University Hospital Heidelberg published a single institution study in the June 1, 2012 European Respiratory Journal (abstract) analyzing the clonal relationship between multifocal NSCLC with indistinguishable morphology in a series of 78 patients.  Their analysis included LOH analysis of 14 polymorphic microsatellite markers and EGFR and KRAS mutation screening using single-strand conformation polymorphism, followed by direct sequencing of abnormally migrating bands. 

This study included 58 adenocarcinomas and 20 squamous cell carcinomas; 59 pT3, 18 pT4, and 1 pM1a. 97.4% of all specimens could be classified as clonal, likely clonal, or non-clonal; only 2 cases were non-informative.

Some results--

1. Tumor nodules of 22 patients (28%) showed divergent LOH status--indicating non-clonal origin. This is a surprising finding--remember this study only included multifocal tumors with indistinguishable morphology.
2. Activating EGFR mutations were found in both synchronous tumors in 4/58 ADC but none of the SQC.
3. KRAS mutations were found in 20 patients (17 ADC, 3 SQC) but in 6 patients the mutation was identified in only 1 of the tumors and in 1 patient there were divergent EGFR and KRAS mutations.
4. With combined allelotyping of LOH and mutational status of EGFR and KRAS, about 36% of cases (28/78 cases) showed evidence that they were not clonally related, including:
5.  14/18 ADC and 6/10 SQC located in the same lobe.
6. Kaplan-Meier curves illustrating the impact of clonality of multifocal NSCLC on overall survival did not identify a statistically significant difference but did show a tantalizing trend that patients with 2 clonally unrelated tumors have a better OS; notably, this difference did reach statistical significance by univariate analysis.

Practical "take-home" points.

There is no specific anatomical or morphological criteria that reliably indicate a clonal or nonclonal origin of multifocal NSCLC tumors. The authors reasonably recommend performing predictive molecular testing (EGFR and KRAS mutational analysis) or allelotyping separately in all cases with synchronous tumors to identify clonally related tumors. This study presents some intriguing data regarding the prognostic value of identifying nonclonalty, but the prognostic impact of clonality needs to be further analyzed in larger cohorts of NSCLC patients with multifocal synchronous tumors.

Establishing clonality in multifocal nonsmall cell lung cancer

Up to 8% of nonsmall cell lung cancer (NSCLC) patients present with 2 or more anatomically separate lung tumors.  Distinguishing between synchronous primary tumors and intrapulmonary metastases is an important clinical question because this determination significantly changes tumor staging and subsequent therapeutic decisions.

Arne Warth et al. have published a single institution study from University Hospital Heidelberg in the current issue (1 June 2012) of European Respiratory Journal (abstract) that examines the clonal relationship in a series of patients with morphologically indistinguishable multifocal NSCLC.

The authors screened their archives for synchronous multifocal NSCLC in the same lobe (pT3), different ipsilateral lobes (pT4), and bilateral lobes (pM1a).  An important part of this study is that the authors only included those cases in which morphology between the tumors was indistinguishable using published criteria from the literature (Girard et al, Am J Surg Pathol 2009--actually a great reference).  They included 78 patients in this study.

The authors performed EGFR and KRAS mutation screening by using single-strand conformation polymorphism sensitive gel elctrophoresis and then subjected aberrently migrating bands to direct DNA Sanger sequencing.  The authors analyzed 14 polymorphic microsatellite markers for loss of heterozygosity. A clever part of the analysis of clonality in this study was the utilization of individual clonality scores for each tumor pair by multiplying the likelihood of the observed findings for each informative variable.  Thus, the clonality score defines the statistical error probability of a clonal or non-clonal relationship.

Results

1. The study included 58 adenocarcinomas (ADC) and 20 squamous cell carcinomas (SQC).  59 were pT3, 18 pT4, and 1 pM1a.  The mean follow up for all patients was 25.6 months. 97.4% of all specimens could be classified as clonal, likely clonal or nonclonal.
2. Tumors from 22 patients (28%) showed divergent LOH status indicative of non-clonal origin (14/58 ADC, 8/20 SQC).
3. Activating EGFR mutations were found in both synchronous tumors in 4/58 ADC and none of the SQC showed EGFR mutation.
4. KRAS mutations were found in 20 cases (17 ADC, 3 SQC), including in both synchronous tumors in 13 cases but in only 1 tumor in 6 cases. 1 patient had divergent EGFR and KRAS mutations in separate tumors.
5. Combined allelotyping of LOH and EGFR and KRAS mutational status showed that 36% of synchronous primary tumors were not clonally related.  Remember these were tumors that were determined to be morphologically indistinguishable.
6. Impact of clonality of multifocal NSCLC on overall survival: Kaplan-Meier curves show a trend that patients with 2 clonally unrelated tumors have a better outcome but this did not reach statistical significance by univariate analysis.

Conculsions

There is no objective, specific anatomical or morphological criteria that reliably indicate a clonal or non-clonal origin of multiple NSCLC tumors.  Therefore, the authors recommend performing predictive molecular testing (EGFR and KRAS) separately in all cases with synchronous tumor nodules or allelotyping to identifiy clonally related tumors.

This paper will influence the way I approach these situations.  First, I will specifically incorporate the Girard criteria to identify morphologically indistinguishable cases.  Second, I will send out separate tumor specimens for EGFR and KRAS mutation analysis before assigning my pathologic staging to the case.  A couple practical questions are not addressed by this study.  Given the remarkable morphological heterogeneity of lung ADC--and the high percentage of "mixed" subtype tumors, isn't it possible that morphological disparate tumors could still be clonally related?  The authors don't specifically address the issue of mixed subtype in relation to "morphologically indistinguishable" tumors--but this would be a major practical issue.  Should we simply assume that morphologically different-ish tumors are non-clonal or do the testing anyway?  A great complimentary study would test all synchronous tumors regardless of histology since this study showed that morphological criteria could not predict clonal origin even in a narrow series of synchronous tumors that looked alike in pairs.  An intriguing point of this study is the possible relationship between clonal relationship and OS.  The authors rightly conclude that this should be tested in larger cohorts of NSCLC patients to identify any statistically significance to the trend that they identified.  I am perplexed as to why clonally unrelated tumors should have better outcomes.  If any one has any thoughts, I'd love to hear them--please post a comment with your thoughts. 

The new IASLC/ATS/ERS Lung Adencarcinoma Classification is a Stage-Independent Predictor of Survival

 I blogged on this new classification over a year ago when it was first published, but now papers are rolling out showing its utility.  My take on this recent paper below is that it confirms the idea of histologically grading lung ADC according to the predominant different subtypes (lepidic predominant grade 1 "well-differentiated" versus solid predominant being grade 3 "poorly-differentiated".  Moreover, it asserts that it is sufficient to regard only the predominant histologic type with regard to survival--which is a relief from going through the tedious task of estimating percentages of different minor subtypes after complete histologic examination.  Get this article--good for discussion with surgeons, oncologists, tumor conference, etc. 

The Novel Histologic International Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society Classification System of Lung Adenocarcinoma Is a Stage-Independent Predictor of Survival.