we collected tumor samples from 372 patients who underwent curative surgical resection for nsclc at seoul national university bundang hospital in korea , between may 2003 and december 2008 . patients who did not undergo curative resection and those who had a history of malignancy , preoperative chemotherapy , or radiotherapy were excluded . smoking status was defined as never - smoker ( < 100 lifetime cigarettes ) or smoker . tumors were staged using the american joint committee on cancer ( ajcc ) tnm classification of malignant tumors seventh edition criteria , and the histological type and grade of differentiation of tumors were determined according to the classification system developed by the world health organization fourth edition . overall survival ( os ) was measured from the date of lung cancer surgery until the time of death , and progression - free survival ( pfs ) was measured from the date of surgery until recurrence or death . all patients provided written informed consent , and this study was approved by the institutional review board ( irb ) of seoul national university bundang hospital . formalin - fixed paraffin - embedded tissues were sectioned at a thickness of 4 m and stained with rabbit monoclonal antibody against human igf1r ( g11 , ventana medical systems , tucson , az , usa ) using an automated immunostainer ( ventana medical systems ) . placental tissue was used as a positive control , and non - immune serum was used as a negative control instead of the primary antibody . adjacent normal - appearing bronchial epithelium within each tissue section served as an internal reference . the h - score ( semiquantitative system with a total score range of 0300 ) was used to evaluate immmunohistochemical staining , and membranous and cytoplasmic staining were evaluated separately . the percentage of positive cells ( 0%100% ) was multiplied by the dominant intensity score of staining . the intensity score was defined as follows : 0 , no appreciable staining ; 1 , barely detectable staining ; 2 , distinct brown staining ; and 3 , strong dark brown staining . a total score range of 0 to 300 was generated for each sample , where median score was used as a cutoff value . all of the slides were evaluated by two pathologists ( e.p . and h.k . ) in a blinded manner . samples were analyzed for egfr mutations within exons 18 to 21 and kras exon 2 ( codons 12 , 13 , and 61 ) using polymerase chain reaction and direct dna sequencing or pyrosequencing methods . in situ hybridization was performed for alk and ros1 rearrangement and met amplification [ 18 - 20 ] . all of the values were based on two - sided statistical analysis , and a p - value of < 0.05 was determined to indicate a statistically significant difference . we collected tumor samples from 372 patients who underwent curative surgical resection for nsclc at seoul national university bundang hospital in korea , between may 2003 and december 2008 . patients who did not undergo curative resection and those who had a history of malignancy , preoperative chemotherapy , or radiotherapy were excluded . smoking status was defined as never - smoker ( < 100 lifetime cigarettes ) or smoker . tumors were staged using the american joint committee on cancer ( ajcc ) tnm classification of malignant tumors seventh edition criteria , and the histological type and grade of differentiation of tumors were determined according to the classification system developed by the world health organization fourth edition . overall survival ( os ) was measured from the date of lung cancer surgery until the time of death , and progression - free survival ( pfs ) was measured from the date of surgery until recurrence or death . all patients provided written informed consent , and this study was approved by the institutional review board ( irb ) of seoul national university bundang hospital . formalin - fixed paraffin - embedded tissues were sectioned at a thickness of 4 m and stained with rabbit monoclonal antibody against human igf1r ( g11 , ventana medical systems , tucson , az , usa ) using an automated immunostainer ( ventana medical systems ) . placental tissue was used as a positive control , and non - immune serum was used as a negative control instead of the primary antibody . adjacent normal - appearing bronchial epithelium within each tissue section served as an internal reference . the h - score ( semiquantitative system with a total score range of 0300 ) was used to evaluate immmunohistochemical staining , and membranous and cytoplasmic staining were evaluated separately . the percentage of positive cells ( 0%100% ) was multiplied by the dominant intensity score of staining . the intensity score was defined as follows : 0 , no appreciable staining ; 1 , barely detectable staining ; 2 , distinct brown staining ; and 3 , strong dark brown staining . a total score range of 0 to 300 was generated for each sample , where median score was used as a cutoff value . all of the slides were evaluated by two pathologists ( e.p . and h.k . ) in a blinded manner . samples were analyzed for egfr mutations within exons 18 to 21 and kras exon 2 ( codons 12 , 13 , and 61 ) using polymerase chain reaction and direct dna sequencing or pyrosequencing methods . in situ hybridization was performed for alk and ros1 rearrangement and met amplification [ 18 - 20 ] . statistical analyses were performed using ibm spss ver . 19.0 ( spss inc . , chicago , il , usa ) . all of the values were based on two - sided statistical analysis , and a p - value of < 0.05 was determined to indicate a statistically significant difference . in non - neoplastic lung tissue , the distribution of igf1r expression was different among cell types . the prevalence of igf1r expression was significantly higher in squamous cell carcinoma ( sqcc ) compared with adc ( p = .000 ) , and representative examples are shown in fig . membranous igf1r expression was identified in 100 of 146 patients ( 68.5% ) and was significantly associated with absence of vascular invasion ( p = .039 ) , absence of lymphatic invasion ( p = .047 ) , absence of perineural invasion ( p = .027 ) , lower stage ( p = .011 ) , and better pfs ( p = .011 ) and os ( p = .034 ) compared to the negative subgroup ( table 2 , fig . multivariate analysis indicated that membranous igf1r expression was an independent prognostic factor for better pfs ( hazard ratio [ hr ] , 0.586 ; 95% confidence interval [ ci ] , 0.352 to 0.975 ; p = .040 ) but not os ( hr , 0.550 ; 95% ci , 0.302 to 1.002 ; p = .051 ) . cytoplasmic igf1r expression was identified in 107 of 146 patients ( 73.3% ) and tended to correlate with younger age ( p = .021 ) and absence of pleural invasion ( p = .013 ) . patients were additionally divided into four groups : group i that was positive for both membranous and cytoplasmic expression ( m+/c+ ) , group ii that was membranous expression - positive and cytoplasmic expression - negative ( m+/c ) , group iii that was membranous expression - negative and cytoplasmic expression - positive ( m/c+ ) , and group iv that was negative for both membranous and cytoplasmic expression ( m/c ) . on univariate analysis , group ii that was membranous expression - positive and cytoplasmic expression - negative ( m+/c ) showed the best pfs ( p = .021 ) and os ( p = .028 ) , followed by group i ( m+/c+ ) , group iv ( m/c ) , and group iii ( m/c+ ) . in adc patients , positive membranous igf1r expression was identified in 31 of 226 patients ( 13.7% ) and was significantly associated with perineural invasion ( p = .040 ) , although the measure had no significant prognostic value . cytoplasmic expression was identified in 141 of 226 patients ( 62.4% ) and correlated with smaller tumor size ( p = .002 ) and absence of vascular invasion ( p = .046 ) , but these results were not statistically significant for pfs or os . the most common mutation was in - frame deletion in exon 19 ( 57 of 108 , 52.8% ) , followed by missense mutation ( l858r ) in exon 21 ( 35 of 108 , 32.4% ) . when adc samples were stratified into egfr - mutant and egfr - wild type subgroups , membranous igf1r expression was identified in 13 of 108 patients ( 12.0% ) in the egfr - mutant adc subgroup . membranous igf1r expression was associated with lymphatic invasion ( p = .039 ) , perineural invasion ( p = .004 ) , solid predominant histology ( p = .037 ) , and higher cancer stage ( p = .030 ) and showed a significantly worse pfs ( p = .002 ) and os ( p = .040 ) compared with those without membranous expression ( table 3 , fig . multivariate analysis revealed that membranous igf1r expression was an independent prognostic factor for worse pfs ( hr , 2.582 ; 95% ci , 1.265 to 5.271 ; p = .009 ) but not for os ( hr , 1.369 ; 95% ci , 0.558 to 3.359 ; p = .492 ) in patients with egfr - mutant adc . cytoplasmic igf1r expression was not associated with any clinicopathological parameters in pfs or os . in egfr - wild type adc , membranous igf1r expression was identified in 23 of 118 patients ( 19.5% ) and was not correlated with clinicopathological parameters . in contrast , cytoplasmic igf1r expression was observed in 86 patients ( 73.5% ) who showed significantly better pfs ( p = .033 ) and os ( p = .05 ) compared to the negative subgroup . no positive membranous igf1r expression was observed for tki - treated egfr - mutant adc ( n = 36 ) . positive igf1r cytoplasmic expression was seen in 15 of 36 patients ( 41.7% ) but was not associated with pfs or os . in addition , igf1r expression was not correlated with alk , kras , ros1 , or met mutation ( p > .05 ) . in non - neoplastic lung tissue , the distribution of igf1r expression was different among cell types . the prevalence of igf1r expression was significantly higher in squamous cell carcinoma ( sqcc ) compared with adc ( p = .000 ) , and representative examples are shown in fig . 1 . membranous igf1r expression was identified in 100 of 146 patients ( 68.5% ) and was significantly associated with absence of vascular invasion ( p = .039 ) , absence of lymphatic invasion ( p = .047 ) , absence of perineural invasion ( p = .027 ) , lower stage ( p = .011 ) , and better pfs ( p = .011 ) and os ( p = .034 ) compared to the negative subgroup ( table 2 , fig . multivariate analysis indicated that membranous igf1r expression was an independent prognostic factor for better pfs ( hazard ratio [ hr ] , 0.586 ; 95% confidence interval [ ci ] , 0.352 to 0.975 ; p = .040 ) but not os ( hr , 0.550 ; 95% ci , 0.302 to 1.002 ; p = .051 ) . cytoplasmic igf1r expression was identified in 107 of 146 patients ( 73.3% ) and tended to correlate with younger age ( p = .021 ) and absence of pleural invasion ( p = .013 ) . patients were additionally divided into four groups : group i that was positive for both membranous and cytoplasmic expression ( m+/c+ ) , group ii that was membranous expression - positive and cytoplasmic expression - negative ( m+/c ) , group iii that was membranous expression - negative and cytoplasmic expression - positive ( m/c+ ) , and group iv that was negative for both membranous and cytoplasmic expression ( m/c ) . on univariate analysis , group ii that was membranous expression - positive and cytoplasmic expression - negative ( m+/c ) showed the best pfs ( p = .021 ) and os ( p = .028 ) , followed by group i ( m+/c+ ) , group iv ( m/c ) , and group iii ( m/c+ ) . positive membranous igf1r expression was identified in 31 of 226 patients ( 13.7% ) and was significantly associated with perineural invasion ( p = .040 ) , although the measure had no significant prognostic value . cytoplasmic expression was identified in 141 of 226 patients ( 62.4% ) and correlated with smaller tumor size ( p = .002 ) and absence of vascular invasion ( p = .046 ) , but these results were not statistically significant for pfs or os . the most common mutation was in - frame deletion in exon 19 ( 57 of 108 , 52.8% ) , followed by missense mutation ( l858r ) in exon 21 ( 35 of 108 , 32.4% ) . when adc samples were stratified into egfr - mutant and egfr - wild type subgroups , membranous igf1r expression was identified in 13 of 108 patients ( 12.0% ) in the egfr - mutant adc subgroup . membranous igf1r expression was associated with lymphatic invasion ( p = .039 ) , perineural invasion ( p = .004 ) , solid predominant histology ( p = .037 ) , and higher cancer stage ( p = .030 ) and showed a significantly worse pfs ( p = .002 ) and os ( p = .040 ) compared with those without membranous expression ( table 3 , fig . multivariate analysis revealed that membranous igf1r expression was an independent prognostic factor for worse pfs ( hr , 2.582 ; 95% ci , 1.265 to 5.271 ; p = .009 ) but not for os ( hr , 1.369 ; 95% ci , 0.558 to 3.359 ; p = .492 ) in patients with egfr - mutant adc . cytoplasmic igf1r expression was not associated with any clinicopathological parameters in pfs or os . in egfr - wild type adc , membranous igf1r expression was identified in 23 of 118 patients ( 19.5% ) and was not correlated with clinicopathological parameters . in contrast , cytoplasmic igf1r expression was observed in 86 patients ( 73.5% ) who showed significantly better pfs ( p = .033 ) and os ( p = .05 ) compared to the negative subgroup . no positive membranous igf1r expression was observed for tki - treated egfr - mutant adc ( n = 36 ) . positive igf1r cytoplasmic expression was seen in 15 of 36 patients ( 41.7% ) but was not associated with pfs or os . in addition , igf1r expression was not correlated with alk , kras , ros1 , or met mutation ( p > .05 ) . the main findings of this study are as follows : ( 1 ) the prevalence of igf1r expression was significantly higher in sqcc compared with adc ; ( 2 ) in sqcc , membranous igf1r expression was associated with significantly better pfs compared with those without membranous expression ; ( 3 ) in adc , membranous and cytoplasmic igf1r expression had no significant prognostic value ; ( 4 ) in the egfr - mutant adc subgroup , membranous igf1r expression was significantly associated with worse pfs . in sqcc , membranous igf1r expression was associated with several good prognostic clinicopathological parameters and showed a linear relationship with pfs . several previous studies have delineated the association between igf1r expression and better os , although the biologic mechanism is unclear . one possible explanation is that , because igf1r is expressed in normal bronchial basal cells , the progenitors of sqcc tumor cells , membranous igf1r expression on tumor cells could be interpreted as an activated and overexpressed form of the physiological function of the receptor . if the tumor cell displays higher igf1r expression , it is a more mature and well - differentiated cell that still depends on the normal igf1r pathway and can be expected to become indolent during the course of the disease . an alternative explanation , based on the fact that the degree of receptor expression would reflect the level of its ligands , is that , if the level of the ligand is higher , more receptors would be occupied and internalized for signaling , while low circulating ligand level would result in receptor overexpression as a compensatory mechanism . therefore , positive membranous igf1r expression could be interpreted as a level of igf1 or igf2 within or lower than the normal range , which could imply that the tumor has low proliferative activity . not only was membranous igf1r expression significantly associated with better survival in sqcc , but the membranous - positive and cytoplasmic - negative subgroup ( m+/c ) had the best pfs and os in sqcc . this is the first study to evaluate the cellular location of igf1r expression in nsclc and provides better understanding of the relevance of pattern of igf1r staining in nsclc . in adc patients , however , when we classified these patients on the basis of egfr - mutation status , the egfr - mutant adc subgroup showed strong correlations with several negative prognostic clinicopathological parameters and worse survival . therefore , igf1r was identified as a poor prognostic factor in egfr - mutant adc , which is opposite the result seen in the sqcc group . this difference prompted speculation that the role of the igf1r pathway in adc might differ according to the presence of egfr mutations . we assume that , in the egfr - mutant adc subgroup , the igf1r pathway would activate downstream targets on the egfr , which could promote cellular proliferation , inhibition of apoptosis , angiogenesis , and anchorage - independent growth . in nsclc , experimental studies have demonstrated that igf1r activation can underlie resistance to egfr - targeted therapies , and several clinical trials have indicated that coexpression of egfr and igf1r correlates with poor survival . furthermore , yeo et al .   recently reported that igf1r expression was a negative predictive factor for response to egfr - tki treatment in egfr - mutant adc patients . although these data indicate that igf1r leads to acquired resistance against egfr - targeted drugs , none of the patients in the egfr - tki treated group showed membranous igf1r expression . in summary , the implication of igf1r expression in nsclc differs according to subtype , and the clinical significance of igf1r expression should be interpreted after considering the histology and genotype of the tumor . high membranous igf1r expression might be a biomarker of better pfs in sqcc and of poor pfs in egfr - mutant lung adc . these findings will serve to improve study design for subsequent investigations into igf1r and nsclc and to select patients for future anti - igf1r therapy .
background : insulin - like growth factor-1 receptor ( igf1r ) is a membrane receptor - type tyrosine kinase that has attracted considerable attention as a potential therapeutic target , although its clinical significance in non - small cell lung cancer ( nsclc ) is controversial . 
 this study aimed to clarify the clinical significance of igf1r expression in human nsclc.methods:igf1r protein expression was evaluated using immunohistochemistry in 372 patients with nsclc who underwent curative surgical resection ( 146 squamous cell carcinomas [ sqccs ] and 226 adenocarcinomas [ adcs ] ) . 
 we then analyzed correlations between expression of igf1r and clinicopathological and molecular features and prognostic significance.results:membranous and cytoplasmic igf1r expression were significantly higher in sqccs than in adcs . in patients with sqcc , membranous igf1r expression 
 was associated with absence of vascular , lymphatic , and perineural invasion ; lower stage ; and better progression - free survival ( pfs ) ( hazard ratio [ hr ] , 0.586 ; p = .040 ) . in patients with adc , igf1r expression did not have a significant prognostic value ; however , in the subgroup of epidermal growth factor receptor ( egfr)-mutant adc , membranous igf1r expression was associated with lymphatic and perineural invasion , solid predominant histology , and higher cancer stage and was significantly associated with worse pfs ( hr , 2.582 ; p = .009).conclusions : lung adc and sqcc showed distinct igf1r expression profiles that demonstrated prognostic significance . 
 high membranous igf1r expression was predictive of poor pfs in egfr - mutant lung adc , while it was predictive of better pfs in sqcc . 
 these findings will help improve study design for subsequent investigations and select patients for future anti - igf1r therapy .