Clinicopathological analysis of Large Cell Lung Carcinomas definitely diagnosed according to the New World Health Organization Criteria

Accepted Manuscript Title: Clinicopathological analysis of Large Cell Lung Carcinomas definitely diagnosed according to the New World Health Organization Criteria Authors: Yalan Bi, Yang Qu, Zhiyong Liang, Zichen Liu, Hui Zhang, Xiaolong Liang, Yufeng Luo, Jinling Cao, Haiqing Zhang, Ruie Feng PII: DOI: Reference:

S0344-0338(17)31130-5 https://doi.org/10.1016/j.prp.2018.02.006 PRP 51996

To appear in: Received date: Revised date: Accepted date:

8-11-2017 15-1-2018 8-2-2018

Please cite this article as: Yalan Bi, Yang Qu, Zhiyong Liang, Zichen Liu, Hui Zhang, Xiaolong Liang, Yufeng Luo, Jinling Cao, Haiqing Zhang, Ruie Feng, Clinicopathological analysis of Large Cell Lung Carcinomas definitely diagnosed according to the New World Health Organization Criteria, Pathology - Research and Practice https://doi.org/10.1016/j.prp.2018.02.006 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

Clinicopathological analysis of Large Cell Lung Carcinomas definitely diagnosed according to the New World Health Organization Criteria

IP T

Yalan Bi1,#, Yang Qu2,#, Zhiyong Liang1, Zichen Liu2, Hui Zhang1, Xiaolong Liang1, Yufeng

SC R

Luo1, Jinling Cao1, Haiqing Zhang2,*, Ruie Feng1,*

1

Department of Pathology, Peking Union Medical College Hospital, Chinese Academy of

U

Medical Sciences and Peking Union Medical College, Beijing 100730, China; 2

N

Department of Pathology, Beijing Chest Hospital, Capital Medical University, Beijing

M

A

101149, China

ED

Running title: Clinicopathological analysis of LCC

Correspondence Author

PT

Ruie Feng1, Haiqing Zhang2 1

CC E

Department of Pathology, Peking Union Medical College Hospital, Chinese Academy of

Medical Sciences and Peking Union Medical College, Beijing 100730, China; 2

A

Department of Pathology, Beijing Chest Hospital, Capital Medical University, Beijing

101149, China

Tel: +86-010-89509381, +86-010-69159375

Fax: +86-010-89509381, +86-010-69159375 1

E-mail: [email protected]; [email protected]

A

CC E

PT

ED

M

A

N

U

SC R

IP T

Note: #Yalan Bi and Yang Qu have contributed equally to this work

2

Abstract

Objective: The definition of large cell lung carcinoma (LCC) has undergone an extensive modification in the World Health Organization (WHO) Classification (2015). Present study aimed to investigate the clinicopathological characteristics of patients diagnosed as LCC

IP T

according to current WHO criteria. Methods: LCCs diagnosed based on the previous WHO

SC R

classification were reevaluated, and 17 cases of LCC were finally identified at Peking Union

Medical College Hospital and Beijing Chest Hospital between 2009 and 2015. The clinicopathologic features were examined and EGFR and KRAS mutations were tested. Survival

U

of the patients was analyzed by Kaplan-Meier method. Results: The median age of the patients

N

was 64 years (range: 40-78). Most patients were male (64.7%) and about half of the patients

M

A

were at TNM stage III (47.1%). Morphologically, most cases (70.6%) were classic LCC. All patients were treated by lobectomy plus lymph node dissection, 2 with bi-lobectomy and 1 with

ED

complex lobectomy, and the other 2 patients were further treated by partial pericardiotomy. Ten

PT

patients received postoperative chemotherapy, while only 3 patients were treated with radiotherapy after surgery. Molecular analysis showed two cases of EGFR mutation (L858R)

CC E

but without non-overlapping KRAS mutation. The 3-year overall survival rate was 48.4±15.1%. Chemotherapy was the only predictive factor that is associated with the prognosis of the patients

A

(P=0.003). Conclusion: The clinicopathological characteristics of 17 cases of stringently diagnosed LCC were retrospectively analyzed. LCC in our study showed aggressive behavior with high recurrence and metastasis and poor prognosis. Chemotherapy was only predictive factor that is significantly associated with the prognosis of the patients. Future studies based on a larger series and long term follow-up are still needed to characterize it further. 3

Keywords: Large cell lung carcinoma (LCC); World Health Organization (WHO) 2015 Classification; Clinicopathological characteristics; Survival; Chemotherapy; EGFR mutation

1 Introduction

IP T

Lung cancer is the most common cause of cancer-related death worldwide [1]. As the most common histological subtype, non-small cell lung carcinomas (NSCLCs) have been reported

SC R

to account for about 80% of lung cancer and rank as the first death cause of malignancy in

China [2, 3]. Large cell lung carcinoma (LCC) is a highly aggressive malignancy that generally considered as the most poorly differentiated type of NSCLCs, accounting for 3% to 9% of all

N

U

cases [4, 5]. In China, the incidence rate of LCC has been reported to be 1% to 5.7% [6].

A

However, it has long been defined as NSCLCs that lack cytologic and architectural features of

M

small cell carcinoma, glandular or squamous differentiation based on routine histologic criteria, according to the World Health Organization (WHO) classification of lung tumors (2004) [7].

ED

That left a large number of NSCLCs lacking specific histologic features to the category of LCC,

PT

which has been suggested to be the main controversial issue in lung cancer classification [8]. The concept of LCC has often been described as a “waste basket” category [9]. Recently,

CC E

increasing attention has focused on immunostaining protocol for assistant diagnosis of specific cell types of LCC [10, 11]. That resulted in the publication of the 4th edition of the WHO

A

Classification of Tumours of the Lung, Pleura, Thymus and Heart in April 2015 [12]. The latest WHO classification (2015) extensively modified the LCC category by approval of immunohistochemistry for assistant diagnosis. The NSCLCs previously classified as LCC are now reclassified according to their immunophenotype for adenocarcinoma markers (most notably TTF1) or squamous cell carcinoma markers (most notably p40). Besides, the 4

uncommon specific cell types, like basaloid carcinoma, large cell neuroendocrine carcinoma, and lymphoepithelioma like carcinoma, are no longer classified under LCC. That resulted in a small number of NSCLCs that diagnosed as LCC, accounting for only 1% to 2% of lung cancers [13]. This category of patients has been reported to have an inferior prognosis [10]. However,

IP T

to our knowledge, the clinical characteristics of this newly defined category have not been well described. In this study, we retrospectively analyzed the clinicopathological characteristics of

SC R

the patients meeting the 2015 WHO criteria for LCC.

2 Material and Methods

N

U

Surgical pathology blocks of LCC based on the previous WHO classification (2004) were

A

reevaluated according to the current WHO criteria (2015). A total of 17 LCC were finally

M

identified from Peking Union Medical College Hospital and Beijing Chest Hospital during 2009 to 2015. Routine hematoxylin-eosin stain was performed for histologic diagnosis. The TNM

ED

edition 8th was used for staging in this study [14]. Patients were treated according to

PT

NCCN Guidelines with the consent of the patients and/or their families [15]. Immunohistochemical analysis was repeated to confirm negative immunophenotype using the

CC E

antibodies against TTF1 (clone 8G7G3/1; 1:100; Maixin, Fujian, China), napsin A (Maixin, Fujian, China), p40 (Maixin, Fujian, China), p63 (clone 4A4; 1:100; Maixin, Fujian, China),

A

and CK5/6 (clone D5/16B4; 1:100; Maixin, Fujian, China). Tumors positive for TTF1 or napsin A were reclassified as adenocarcinoma, while ones diffusely positive for p40, p63 or CK5/6 were reclassified as squamous cell carcinoma. Clinical data, including age, sex, smoking history, tumor size and location, lympho node metastasis at surgery, treatment strategies

5

(surgery, chemotherapy, and/or radiotherapy), and survival status of the patients, were obtained from clinical chart review of each patient.

Molecular analysis of EGFR and KRAS mutations was carried out using amplification refractory mutation system (ARMS) assay [16]. Briefly, DNA was extracted from 4-5 unstained

IP T

4 μm thick sections of tissue with tumor cellularity > 75% using a DNA extraction kit (Amoy

SC R

Diagnostics Co., Ltd, Xiamen, China) according to the manufacturer's instructions. DNA concentration was quantified by ultraviolet spectrophotometry (Amoy Diagnostics Co., Ltd,

Xiamen, China). ARMS was performed using ADx-EG01 EGFR and ADx-KR01 KRAS

U

Mutation Test Kits (Amoy Diagnostics Co., Ltd, Xiamen, China). Mutations of EGFR and

N

KRAS were examined on the Cobas z 480 Analyzer (Roche, Basel, Switzerland) using extracted

M

A

DNA (42.3μL EGFR, 45μL KRAS, 2μg/μL). The PCR program was: 94˚C for 5 min, 15 cycles at 95˚C for 25 sec, at 64˚C for 20 sec and at 72˚C for 20 sec, and followed by 31 cycles at 93˚C

ED

for 25 sec, at 60˚C for 35 sec and at 72˚C for 20 sec. EGFR mutations were detected in exons

PT

18 to 21, whereas KRAS mutations were detected in exons 12 and 13.

Survival of the patients was calculated from the date of surgery to the death or last observation.

CC E

Data were analyzed using the SPSS 17.0 software program (SPSS, Inc., Chicago, IL, USA). Continuous data were expressed as mean ± standard deviation (SD) and categorical data were

A

as numbers or percentages. Kaplan-Meier method was used for survival analysis and differences were analyzed by log-rank tests. A P value of less than 0.05 was considered statistically significant.

3 Results

6

The study included 17 cases of LCC definitely diagnosed according to the WHO 2015 criteria. Morphologically, most cases showed nests or sheets of cells with neither glandular nor squamous differentiation (Fig. 1). All cases were definitely diagnosed based on the absence of both adenocarcinoma (TTF1 and napsin A) and squamous cell carcinoma markers (p40 and

IP T

p63).

SC R

Clinical characteristics of the patients were summarized in Table 1. The median age of the

patients was 64 years (range: 40-78 years). There was an obvious male predominance with 11 males and 6 females. Seven patients were current smoker, one was former smoker, and the

U

median smoking pack-years was 40 (range: 20–50). The rest 9 patients were nonsmokers. Most

N

tumors (10) were typically peripheral masses, accounting for 58.8% of all cases. The median

M

A

tumor size was 4.0 cm, ranging from 1.6 to 10 cm. Most patients (14/17, 82.4%) had tumor invasiveness (Table 1). Six patients were at TNM stage IIIA, 3 at stage IB, and each 2 patients

ED

were at stage IA, IIA, IIB and IIIB, respectively. Morphologically, 12 cases (70.6 %) were

PT

classic LCC, 2 showed basaloid squamous carcinoma, and each one showed multinucleated giant cells (8 %) and clear cells (20 %). The other one showed clear cells (70 %) with few

CC E

multinucleated giant cells (5 %). Twelve patients underwent lobectomy plus lymph node dissection, 2 patients underwent bi-lobectomy plus lymph node dissection, 2 patients underwent

A

lobectomy plus lymph node dissection and partial pericardiotomy, and the other one underwent complex lobectomy (right middle and part of lower and upper lung lobes) plus lymph node dissection (Table 2). Only one patient was resected by thoracoscopy. Ten patients received postoperative chemotherapy, and the regimens used were taxol plus carboplatin, pemetrexed

7

plus cisplatin, vinorelbine plus carboplatin and etc. Only 3 patients were treated with radiotherapy after surgery.

Prognosis of the patients was analyzed and 3 patients were lost of follow-up. A total of 14 patients were followed up for 36.00 months (range: 7-65 months). Six patients died during the

IP T

follow-up and all died from cancer. Among which, one patient had brain metastasis and each

SC R

two patients had bone and multiple liver metastases, respectively. Only one patient received radiotherapy after disease progression and the remaining 4 patients refused the further

treatment. Among alive patients, one patient showed left temporal lobe lesion, two patients

U

developed metastases (brain and intrapulmonary metastasis, respectively), and all had

N

undergone further radiotherapy. The rest patients were progression free during the follow-up.

M

A

Molecular analysis showed 2 cases of EGFR mutation (L858R, case no. 6 and 12), and both the patients were still alive. However, no case of non-overlapping KRAS mutation was identified

ED

in our study.

PT

Survival analysis showed the 2- and 3-year overall survival rates of 67.7±13.5 % and 48.4±15.1%, respectively (Fig. 2A). Chemotherapy was only predictive factor that was

CC E

associated with the favorable prognosis of the patients (P=0.003, Table 3 and Fig. 2B). Other parameters, including age, gender, smoking status, tumor location and size, TNM stage, lympho

A

node metastasis, pathological type, and radiotherapy, were not found to be significantly associated with prognosis of the patients.

4 Discussion

8

According to the new 2015 WHO classification, LCC is defined as an undifferentiated NSCLCs that lacks the cytological, architectural and immnuohistochemical features of small cell carcinoma, adenocarcinoma, or squamous cell carcinoma [12]. However, to our knowledge, few studies have investigated the clinical characteristics of this newly-defined category. The

IP T

present study mainly focused on the clinicopathological features and prognosis of LCC patients diagnosed according to the latest WHO criteria (2015). In our study, 17 cases of definitely

SC R

diagnosed LCC were retrospectively analyzed. The median age of our patients was 64 years,

and most patients were male (64.7%). However, fewer cases of smoker were observed in our

U

study. These data were partially consistent with the results reported by previous study, which

N

indicated a median age of 62 years, and a male and smoking proportion of 55.0% and 95.0%,

A

respectively [10], corroborating the description of LCC given by the 2015 WHO criteria.

M

However, in the study by Brandon et al, a male: female ratio of 1:1 was obtained, since only 2

ED

patients were finally reclassified to LCC during 1974 to 1991 according to the 2015 WHO criteria [17]. In our study, 8 patients were at TNM stage III (47.1%), followed by 5 patients at

PT

stage I (29.4%), and the other 4 patients were at stage II. Among the 20 patients classified as

CC E

marker-null LCC (LCC-null) reported by Rekhtman et al, most patients were at early stages (stage I, 40%), and each 6 patients were at stage II and III/IV (30%) [10]. According to the new WHO classification (2015), LCC are typical peripheral masses. Our study here showed that

A

more cases of the LCC were of a peripheral type (10/17, 58.8%), while about 41.2% of cases were central type LCC. The clear cell and rhabdoid phenotypes have been on longer considered as subtypes of LCC, and rhabdoid cytology needs to be documented as a percentage of the total tumor that is prognostically significant in 2015 WHO criteria. Most tumors were classic LCC, 9

not otherwise specified (NOS), accounting for 70.6 % of all cases, consistent with the rate (65%) reported by Rekhtman et al [10].

Mutation profiles of immunophenotype-based LCC have been determined previously. EGFR and KRAS mutations have long been identified in lung cancer patients. EGFR mutation has

IP T

been reported to be more frequent in female, non-smoker and Asian population [18-20]. It has

SC R

been reported in about 10% of the Western population and in about 50% of Asian lung cancer

patients [21]. However, unlike the EGFR mutation, KRAS mutation showed no sex predilection, and was suggested to be more frequent in white populations and former or current smokers [22,

U

23]. Molecular analysis of EGFR and KRAS mutations in our study showed that, among the 17

N

cases of well-definited LCC, 2 cases had EGFR mutation (L858R) andno case of non-

M

A

overlapping KRAS mutation was detected. All two cases with EGFR mutation were female and never smokers. LCC-N previously defined as negative for adenocarcinoma marker, squamous

ED

cell carcinoma marker, and mucin were found to have KRAS mutation (G12C) [24]. Five cases

PT

of KRAS mutation have been reported by Rekhtman and colleagues in a set of 20 LCC-null before publication of the new WHO classification (2015), while no EGFR mutation was

CC E

identified in their study [10]. Moreover, the study by Driver and colleagues reported that one of two case of LCC reclassified according to the WHO 2015 classification had KRAS mutation

A

(G12C), and there was still no case of EGFR mutation reported [17]. These results may be in favor of ethnic difference in EGFR and KRAS mutations between Western and Asian population, although the sample sizes involved were small.

Previous study by Rekhtman and colleagues indicated that patients with LCC-null had a distinctly inferior prognosis when compared with their marker-positive counterparts. The 510

year overall survival rate of 12% was observed in their LCC-null patients [10]. In the present study, a 3-year overall survival rate of 48.4±15.1% was obtained. Among the 14 LCC patients with complete follow-up, 8 patients 47.1%) developed recurrence (Left temporal lobe) and/or distant metastasis (multiple live, brain, bone, and intrapulmonary metastasis), suggesting a high

IP T

risk of recurrence and metastasis of the LCC. Log-rank analysis showed that the use of chemotherapy was significantly associated with prognosis of our patients. However, in view of

SC R

the small sample size involved in our study, multivariate analysis was not performed to adjust

for other factors. Besides, the follow-up time in our study was comparatively shorter. Moreover,

U

some of our patients might not be properly managed (according the NCCN guideline) due to

N

the complex clinical condition. Future perspective study based on a large population of

A

definitely diagnosed LCC with longer follow-up are therefore needed to further clarify the

M

prognosis of the LCC patients and identify the potential factors for better prognosis prediction.

ED

In conclusion, we retrospectively analyzed the clinicopathological characteristics of 17

PT

stringently diagnosed LCC based on the current WHO criteria (2015). LCC in our study showed aggressive behavior with high recurrence and metastasis and poor prognosis. Chemotherapy

CC E

was only predictive factor that significantly associated with the survival of the patients. Future studies based on a larger series and long term follow-up should be performed to clarify it

A

further.

Compliance with Ethical Standards

11

The study was performed according to the Declaration of Helsinki and ethics approval was not required due to the retrospective nature of the study. Informed consents were obtained from all participants.

IP T

Conflict of interest

The authors declare that they have no conflict of interest.

SC R

Acknowledgement

This work was supported by the Central Public-interest Scientific Institution Basal Research

U

Fund (no.A463800).

A

Ferlay J, Soerjomataram I, Ervik M, Dikshit R, Eser S, Mathers C, Rebelo M, Parkin

M

1.

N

Reference

D, Forman D, Bray F: Cancer Incidence and Mortality Worldwide: IARC CancerBase

ED

No. 11 [Internet]. Lyon, France: International Agency for Research on Cancer.

2.

PT

GLOBOCAN. 2013; 2012 v1. 0, htt p. globocan iarc fr Accessed 30(2015). Fukuoka M, Wu Y-L, Thongprasert S, Sunpaweravong P, Leong S-S, Sriuranpong V,

CC E

Chao T-Y, Nakagawa K, Chu D-T, Saijo N: Biomarker analyses and final overall survival results from a phase III, randomized, open-label, first-line study of gefitinib

A

versus carboplatin/paclitaxel in clinically selected patients with advanced non–small-

3.

cell lung cancer in Asia (IPASS). J Clin Oncol 29(2011):2866-2874. Huang J-J, Ma S-X, Hou X, Wang Z, Zeng Y-D, Qin T, Dinglin X-X, Chen L-K: Toxic epidermal necrolysis related to AP (pemetrexed plus cisplatin) and gefitinib combination therapy in a patient with metastatic non-small cell lung cancer. Chin J 12

Cancer 34(2015):94. 4.

Bethesda M: based on November 2010 SEER data submission, posted to the SEER web site. (2011).

5.

Lag R, Harkins D, Krapcho M, Mariotto A, Miller B, Feuer E, Clegg L, Eisner M,

IP T

Horner M, Howlader N: Seer cancer statistics review. Bethesda, National Cancer Institute (1975):1975-2003.

Liang R, Chen TX, Wang ZQ, Jin KW, Zhang LY, Yan QN, Zhang HH, Wang WP: A

SC R

6.

retrospective analysis of the clinicopathological characteristics of large cell carcinoma

Travis WD, Harris C: Pathology and genetics of tumours of the lung, pleura, thymus

N

7.

U

of the lung. Exp Ther Med 9(2015):197-202.

Pelosi G, Barbareschi M, Cavazza A, Graziano P, Rossi G, Papotti M: Large cell

M

8.

A

and heart: Feance: IARC Press, 2004; 2004.

ED

carcinoma of the lung: a tumor in search of an author. A clinically oriented critical reappraisal. Lung Cancer 87(2015):226-231. Kerr KM: Pulmonary Pathology: SY22-1 UPDATE ON LARGE CELL

PT

9.

CC E

CARCINOMAS OF THE LUNG. Pathology-Journal of the RCPA 46(2014):S39. 10.

Rekhtman N, Tafe LJ, Chaft JE, Wang L, Arcila ME, Colanta A, Moreira AL, Zakowski

A

MF, Travis WD, Sima CS: Distinct profile of driver mutations and clinical features in immunomarker-defined subsets of pulmonary large-cell carcinoma. Mod Pathol 26(2013):511-522.

11.

Rossi G, Mengoli M, Cavazza A, Nicoli D, Barbareschi M, Cantaloni C, Papotti M, Tironi A, Graziano P, Paci M: Large cell carcinoma of the lung: clinically oriented 13

classification integrating immunohistochemistry and molecular biology. Virchows Arch 464(2014):61-68. 12.

Travis WD, Brambilla E, Burke A, Marx A, Nicholson AG, Cancer IAfRo: WHO classification of tumours of the lung, pleura, thymus and heart: International Agency

13.

IP T

for Research on Cancer; 2015. Lewis DR, Check DP, Caporaso NE, Travis WD, Devesa SS: US lung cancer trends by

14.

SC R

histologic type. Cancer 120(2014):2883-2892.

Detterbeck FC, Boffa DJ, Kim AW, Tanoue LT: The Eighth Edition Lung Cancer Stage

Ettinger D, Borghaei H, Akerley W: NCCN Clinical Practice Guidelines in Oncology

N

15.

U

Classification. Chest 151(2017):193.

Li X, Wang X, Zhu H, Liu C, Zhou X, Zhao B, Duan H, Yang J, Gu G, Zhan Y:

ED

16.

M

Network Jnccn 4(2015):548.

A

– Non-Small Cell Lung Cancer. Journal of the National Comprehensive Cancer

Epidermal growth factor receptor gene mutations in patients with lung adenocarcinoma

PT

differ by frequency and type between Uighur and Han ethnic groups in Xinjiang

CC E

Autonomous Region. BMC Genet 16(2015):1. 17.

Driver B, Portier B, Mody D, Deavers M, Bernicker E, Kim M, Teh B, Santacruz J,

A

Kopas L, Munden R: Next-Generation Sequencing of a Cohort of Pulmonary Large Cell Carcinomas Reclassified by World Health Organization 2015 Criteria. Arch Pathol Lab Med (2015).

18.

Gazdar A: Activating and resistance mutations of EGFR in non-small-cell lung cancer: role in clinical response to EGFR tyrosine kinase inhibitors. Oncogene 28(2009):S2414

S31. 19.

Shigematsu H, Lin L, Takahashi T, Nomura M, Suzuki M, Wistuba II, Fong KM, Lee H, Toyooka S, Shimizu N: Clinical and biological features associated with epidermal growth factor receptor gene mutations in lung cancers. J Natl Cancer Inst

20.

IP T

97(2005):339-346. Dogan S, Shen R, Ang DC, Johnson ML, D'Angelo SP, Paik PK, Brzostowski EB, Riely

SC R

GJ, Kris MG, Zakowski MF: Molecular epidemiology of EGFR and KRAS mutations

in 3,026 lung adenocarcinomas: higher susceptibility of women to smoking-related

Migali C, Bielinska AM, Bhosle J, O'Brien M: Development in the diagnostic lung

N

21.

U

KRAS-mutant cancers. Clinical Cancer Research 18(2012):6169-6177.

A

cancer pathway: implication for treatment. Lung Cancer Management 3(2014):417-

Suzuki M, Shigematsu H, Iizasa T, Hiroshima K, Nakatani Y, Minna JD, Gazdar AF,

ED

22.

M

428.

Fujisawa T: Exclusive mutation in epidermal growth factor receptor gene, HER‐2,

PT

and KRAS, and synchronous methylation of nonsmall cell lung cancer. Cancer

CC E

106(2006):2200-2207. 23.

Buttitta F, Barassi F, Fresu G, Felicioni L, Chella A, Paolizzi D, Lattanzio G, Salvatore

A

S, Camplese PP, Rosini S: Mutational analysis of the HER2 gene in lung tumors from Caucasian patients: mutations are mainly present in adenocarcinomas with bronchioloalveolar features. Int J Cancer 119(2006):2586-2591.

24.

Project TCLCG, NGM NGM: A genomics-based classification of human lung tumors. Sci Transl Med 5(2013):209ra153. 15

I N U SC R

Smoking pack-years* 0

Size (cm) 2.8

Location

LNM

Tumor invasiveness

TNM

Pathology

Male

Age years 51

Center

0/20

NOS

2

Male

64

30

8

Center

Main bronchus, lower lobe obstructive pneumonia Lung pleura, bronchus

IB

IIIA

NOS

3

Male

56

0

2.4

Center

A

Table 1 Clinical characteristic of the patients with large cell lung cancer reclassified according to the WHO 2015 Criteria

1

IIB

NOS

4

Male

56

0

3.5

IB

NOS

5

Female

65

0

6

6

Female

59

0

7

Male

64

40

8

Male

78

50

9

Female

66

10

Male

71

11

Female

70

12

Female

13

0/20

Periphery

0/13

Pleura mediastinalis, part of the pericardium No

Periphery

4/16

Chest wall

IIIB

NOS

M

Gender

ED

No.

0/21

Center

0/9

No

IA

NOS

3

Center

0/53

Lung pleura and bronchus

IB

NOS

4.5

Periphery

4/26

IIIA

NOS

0

3

Periphery

5/16

Visceral pleura, intravascular thrombus Visceral lung pleura

IIIA

50

10

Center

0/30

Lung pleura and bronchus

IIIA

CC (70%)+MGC (5%) NOS

0

5

Periphery

7/27

IIIB

CC (20%)

61

0

4

Periphery

1/30

Pericardium, left phrenic nerve, lung pleura, bronchus Chest wall, lung pleura

IIIA

NOS

Male

40

20

4.2

Periphery

0/48

Lung pleura

IIA

MGC (8%)

14

Male

67

0

5

Periphery

0/43

Bronchus

IIA

BSC

15

Male

72

40

2

Periphery

0/24

No

IA

BSC

16

Male

68

40

1.6

Periphery

1/16

Intravascular thrombus

IIIA

NOS

17

Female

64

40

5.5

Center

0/22

Bronchus

IIB

NOS

A

CC E

PT

3

Note: LNM: Lympho node metastasis; NOS: Not otherwise specified; TGC: Tumor giant cells; CC: Clear cell; MGC: Multinuclear giant cell; BSC: Basaloid squamous carcinoma; Pack-years: Packs of cigarettes smoked per day × years of smoking

16

I N U SC R

Table 2 Clinical management and prognosis of the patients with large cell lung cancer reclassified according to the WHO 2015 Criteria No.

Surgery

PostRT

Post-CT (Cycle)

1

Bi-lobectomy+lymph node dissection

No

TC (4)

2

Lobectomy+lymph node dissection

No

TC (2)

Brain/30

3

No

No

NA

NA

4

Lobectomy+lymph node dissection+partial pericardiotomy Lobectomy+lymph node dissection

No

NA

NA

5

Bi-lobectomy+lymph node dissection

No

PC (2)

6

Lobectomy+lymph node dissection

No

No

Mediastinal lymph No nodes+Brain

7

Lobectomy+lymph node dissection

No

PC (4)

No

Death/24

8

Lobectomy+lymph node dissection

No

P (4)

No

Alive/7

9

Lobectomy+lymph node dissection

No

VC (4)

No

Post-progression therapy

No

A

M

ED

PT

10

No

Progression /months

Whole brain radiotherapy

Whole brain radiotherapy

Status/months Alive/60 Alive/56

Alive/14 Alive/17

Alive/65 #

No

No

Local+Bone/5

No

Death/7

No

VC (4)

Brain/25

No#

Death/31

12

Complex lobectomy (medium and part of lower and upper lobes)+lymph node Lobectomy+lymph node dissection dissection+partial pericardiotomy Lobectomy+lymph node dissection

Yes

DC (4)

Head radiotherapy

Alive/47

13

Lobectomy+lymph node dissection

No

GC (4)

Left temporal lobe/29 No

CC E

11

Alive/46 #

Lobectomy+lymph node dissection

No

No

Multiple liver/12

No

Death/12

15

Lobectomy+lymph node dissection

Yes

PC (4)

Bone/13

Radiotherapy

Death/36

Lobectomy+lymph node dissection*

NA

NA

NA

A

14

16 17

Lobectomy+lymph node dissection

No

No

Multiple liver/3

NA #

No

Death/12

Note: *Thoracoscopy; LNM: Lympho node metastasis; RT: Radiotherapy; CT: Chemotherapy; TC: Taxol+Carboplatin; PC: Pemetrexed+Cisplatin; P: Pemetrexed; VC: Vinorelbine+Carboplatin; DC: Docetaxel+Cisplatin; GC: Gemcitabine+Cisplatin. #The patients refused to receive further treatment.

17

Table 3 Survival analysis of patients with pulmonary large cell carcinoma χ2

P value

Age (≥65years)

2.314

0.128

Sex

0.228

0.633

Smoking

1.067

0.302

Tumor size (>5 cm)

0.664

0.415

Tumor location

0.411

0.522

TNM stage

0.570

Lympho node metastasis

1.009

Pathological type

2.093

IP T

Variable

0.450 0.315

SC R

0.553

Radiotherapy Chemotherapy

0.360

8.997

0.003

0.276

0.599

A

CC E

PT

ED

M

A

N

U

Targeted therapy

0.836

18

Figure legends

Fig. 1 Histopathology analysis of large cell lung carcinoma (LCC). HE staining showed nests or sheets of tumor cells (x100); Immunohistochemical analysis of expressions of

IP T

adenocarcinoma (TTF1 and napsin A) and squamous cell carcinoma markers (p40 and p63).

Fig. 2 Kaplan-Meier survival analysis of patients with large cell carcinoma. (A) Overall

A

CC E

PT

ED

M

A

N

U

SC R

survival; (B) survival of the patients stratified by chemotherapy.

19

A ED

PT

CC E

IP T

SC R

U

N

A

M

Figr-1

20

A ED

PT

CC E

IP T

SC R

U

N

A

M

Figr-2

21