Avances en patología
pulmonar
María Dolores Lozano
Clínica Universidad de Navarra
Barcelona, 11 Junio 2015
Definición, criterios diagnósticos e implicaciones de
STAS en resecciones de NSCLC
Referencias:
Spread through alveolar spaces: A novel pattern of invasion associated with poor
prognosis in 411 small (≤ 2 cm) stage I lung adenocarcinoma . Kyuichi Kadota, Jun-ichi
Nitadori, Camelia Sima, David J Jones, William Travis, Prasad Adsumilli. Memorial
Sloan Cancer Center, New York, NY (Abstract 1924)
Prognostic significance of “spread through alveolar spaces” in mucinous adenocarcinomas
of the lung. Adina Paulk Sandly Liu, Borislav A, Allen Burke. University of Maryland
Medical Center, Baltimore, MD. (Abstract 1954)
Kadota K, Nitadori JI, Sima CS, Ujiie H, Rizk NP, Jones DR, Adudumilli PS, Travis WD. Tumor spread through air spaces
is an important pattern of invasión and impacts the frequency and location of recurrences following limited resection for
small stage I lung adenocarcinomas. J Thorac Oncol 2015;10: 806-14
El concepto “diseminación tumoral a través de espacios alveolares – STAS”
se define como una forma de invasión caracterizada por diseminación de
células tumorales aisladas o grupo tumorales a través de los espacios
alveolares (STAS). Es un patrón de invasión no existente en ningún otro
órgano debido a las características específicas y únicas de la anatomía
pulmonar.
Hay tres patrones morfológicos:
 Micropapilar
 Nidos sólidos
 Células aisladas
Spread through alveolar spaces: A novel pattern of invasion associated with poor
prognosis in 411 small (≤ 2 cn) stage I lung adenocarcinoma (Abstract 1924)
411 resecciones de Ac ≤ 2 cm
resección atípica 120
Lobectomía 291
STAS 155 (38%)
Periódo libre de recurrencia menor (52% vs 80%, p<0,001)
Resección atípica + STAS
Riesgo de metastasis & recidiva locoregional (42,6% vs 10,9%.
P<0,001)
Lobectomía + STAS
No correlación con estos parámetros (p=0,50; p=0,76)
Patrón micropapilar
Células sueltas
Patrón sólido
J Thorac Oncol 2015;10: 806-14
Conclusiones:
The presence of STAS is a significant risk factor of recurrence in
small lung adenocarcinomas treated with limited resection.
These findings support our proposal that STAS should formally
be recognized as a pattern of invasion in lung adenocarcinoma.
J Thorac Oncol 2015;10: 806-14
Prognostic significance of “spread through alveolar spaces” in mucinous
adenocarcinomas of the lung. Adina Paulk Sandly Liu, Borislav A, Allen Burke. University
of Maryland Medical Center, Baltimore, MD. (Abstract 1954)
El significado clínico de STAS en AC mucinosos no está bien estudiado.
30 resecciones: 2 Ac Coloides / 28 Ac mucinosos invasivos
22 STAS
células aisladas, patron sólido, túbulos (15)
patron micropapilar (7)
Conclusiones:
STAS is common in mucinous adenocarcinomas. The micropapillary type
has prognostic significance in terms of metastasis (p=0,03) and recurrence
(p=0,04)
Marcadores de respuesta inmune
en NSCLC (PDL1, PD1 )
PDL-1 es una importante molécula inmunorreguladora que
bloquea la respuesta inmune citotóxica en una variedad de
procesos fisiológicos y patológicos La interacción PD1/PD
L1 resulta en tolerancia inmune frente al tumor
La inhibición PD1/PDL1 reestablece la tolerancia inm
une frente al tumor: Inmunoterapia
Ensayos clínicos en curso muestran resultados prometedores con dianas dirigidas frente a
PDL-1 y su receptor PD-1.
Resultados preliminares de estos ensayos clínicos indican que la sobrexpresión de PDL-1
en las células tumorales mediante inmunohistoquímica puede predecir la respuesta clínica
frente a terapias PD-1/PDL-1.
Comparison of Programmed cell death ligand-1 (PDL-1) expression in main tumor and lymph node
metastasis of stage II and III lung adenocarcinomas. (Abstract 1897)
¿La expresión de PDL-1 cambia con la progresión del tumor y/o es diferente en vari
as localizaciones de un tumor determinado?
Comparar la expresión de PDL-1 en el tumor primario y las metástasis ganglionares
en Ac de pulmón estadios II y III
Examinar la asociación entre expresión de PDL-1 con aspectos clinicopatológicos
109 Ac sin neoadyuvancia
(74 en el abstract)
PDL-1 (clone: E1L3N,Cell Signalling Technology)
Positivo si ≥ 5% de las células tumorales expresan tinción de membrana de cualquie
r
intensidad
Inmunohistoquímica con Ac antiCD8 (4B11,RTU,Leica biosystems)
 PDL1 se asocia con presencia de linfocitos intratumorales CD8+ en el tumor primari
o
 75% de ganglios N1 y N2 muestran la misma expresión de PDL1 que el tumor prim
ario, por tanto la expresión de PDL1 parece no estar alterada con la progresión del t
umor
Clinicopathological and molecular features associated with programmed cell death ligand-1 (PDL-1)
expression in resected lung squamous cell carcinomas. Massachusetts General Hospital, Boston,
MA. (Abstract 1916)
Pocos datos sobre características clinicopatológicas y moleculares de carcinomas
escamosos que expresan PDL-1.
Diseño:
 TMAs de 159 pacientes con Ca. escamosos de pulmón. Estadio I: 94; II: 40;
III: 22; IV: 3
 PDL-1 (clone: E1L3N,Cell Signalling Technology) es positivo si se observa
tinción de membrana en ≥5% de las células tumorales.
 Estudian los TIL con CD8. .(4B11 mouse mAb (leica b)
 Estudio molecular mediante SNaPshot. (153 casos)
Resultados:
 Sobrexpresión de PDL-1 en 42 (26%)
 Mutaciones: PIK3CA: 11; TP53: 11.
 Amplificaciones: FGFR: 15; PDGFR: 4
 Observan relación entre expresión de PDL-1 y presencia de TIL
(p=0,007) y estadios avanzados (I vs II – IV, p=0,0058).
 De 11 tumores con mutaciones en PIK3CA, 7 expresaban PDL-1
(p=0.012)
Conclusiones:
 Sobrexpresión de PDL-1 por IHQ se asocia significativamente a la
presencia de TIL y a estadios avanzados en Carcinomas escamosos
de pulmón resecados
 Existe asociación con mutaciones en PIK3CA
Clinicopathological and molecular features associated with programmed cell death ligand-1 (PDL-1)
expression in resected lung adenocarcinomas. Massachusetts General Hospital, Boston, MA.
(Abstract 1917)
Pocos
datos
sobre
características
clinicopatológicas
y
moleculares
adenocarcinomas de pulmón que expresan PDL-1.
Diseño:
 TMAs de 141* pacientes con Ac de pulmón. Estadio 0: 2; I: 93; II:
23; III: 11; IV: 12
 Patrones: lepídico-48; acinar-49; papilar-16; micropapilar-6; sólido18; variantes-4.
 PDL-1 (clone: E1L3N,Cell Signalling Technology) es positivo si se
observa tinción de membrana en ≥5% de las células tumorales.
 Estudian los TIL con CD8.(4B11 mouse mAb (leica b)
 Estudio molecular mediante SNaPshot.
de
Resultados:
 Sobrexpresión de PDL-1 en (15,7%)
 Mutaciones KRAS: 56; EGFR: 34; otras en 5 casos*
 Observan relación entre expresión de PDL-1 y
 hábito tabáquico (>10 paq/año; p=0,0019),
 patrones sólido (p=0,018) y acinar de alto grado
(p=0,0024)
 presencia de TIL (p=0,014).
 Expresión de PDL-1 se asocia a mutaciones en KRAS (27%,
p=0.0003)
 No hay asociación estadísticamente significativa entre expresión
de PDL-1 y periodo libre de enfermedad (p=0,9)
Conclusiones:
 Sobrexpresión de PDL-1 por IHQ se asocia
significativamente a la presencia de TIL
 Grado nuclear alto, invasión vascular, tamaño del
tumor
 Existe asociación con mutaciones en KRAS
Novedades USCAP en tumores
neuroendocrinos de pulmón
Referencias:
Beyond mitosis and necrosis: Additional histo-Cytologic differences between typical and atypical carcinoids of
the lung and their prognostic significance (Abstract 1963). Lauren Rosen, Ihab Lamzabi, Vijaya Reddy, Paolo
Gattuso. Rush University Medical Center, Chicago, IL.
Architectural differences between Typical and atypical carcinoids of the lung and their prognosis significance.
Ihab Lamzabi et al. Rush University Medical Center, Chicago, IL (Abstract 1929)
Tumores neuroendocrinos bien
diferenciados de pulmón
Grado I: Carcinoide típico (CT)
Mitosis y necrosis
Grado II: Carcinoide atípico (CA)
Hasta 20% fallecen a los 5 años
Objetivo: mejorar el diagnóstico de “subtipos de alto riesgo” de CT y CA
usando criterios histológicos objetivos
Revisión de 50 casos (35 CT y 15 CA). Criterios cito-histológicos asociados a metástasis
Metástasis ganglionares
Metástasis a distancia
Datos
morfológicos
Mitosis
Necrosi
s
Multinuclea
ción
Relación
N/C
Mitosis
Necrosis
Tamaño
Nuclear
Cromati
na
grumos
Patrón
en
nidos
p
0,01
0,008
0,004
0,01
<0,001
<0,001
0,01
0,001
0,001
Revisión de 50 casos (35 CT y 15 CA). Criterios arquitecturales
Patrón en
nidos
Permeación
del pulmón
adyacente
Infiltración
Cápsula
Crec.
trabecular
TC (35)
8/35 (22%)
11/35 (31%)
18/35 (51%)
10/35 (28%)
22/35 (63%)
AC (15)
10/15 (67%)
12/15 (80%)
13/15 (87%)
1/15 (6%)
3/15 (20%)
TC vs AC
P=0,001
P=0,002
P=0,01
P=0,02
P=0,004
Criterios citohistológicos y arquitecturales en CT y CA de significado pronóstico
Mitosis y necrosis (p<0,001)
Metástasis ganglionares y a distancia
Pleomorfismo nuclear
Núcleos grandes
Relación N/C aumentada
Cromatina en grumos gruesos
Patrón en nidos
Permeación del pulmón adyacente
Estratificación de pacientes con CT y CA en categorías de bajo y alto riesgo
Expression of PAK1 and PAK2 is associated with grade of pulmonary neuroendocrine tumors. Stephen Smith, Adam
Bissonnette, David Cohen, Cynthia Timmers, Jin Jen, Junya Fukuoka, Teri Franks, William Travis, David Carbone,
Konstantin Shilo. Ohio State University, Columbus, OH; Mayo Clinic, Rochester, MN; Nagasaki University,
Nagasaki, Japan; Joint Pathology Center, Silver Spring, MD; Memorial Sloan Kettering Cancer Center, New York,
NY. (Abstract 1967)
P21-activated kinasas (PAKs), contribuyen a regular supervivencia celular y proliferación
Sobreexpresión de PAK1 y PAK2 documentada en ca escamoso y papilomas
Objetivo: estudiar al sobreexpresión IHQ de PAK 1 y PAK 2 en varios tipos de tumores
de pulmón (especialmente TNE)
392 tumores usando microarrays de tejidos
Score: negativo, baja expresión, alta expresión (citoplasma)
Resultados y Conclusiones:
Expresión en varios tipos de tumores de pulmón con diferentes frecuencias
PAK1 / tumores carcinoides
PAK2 / neuroendocrinos de alto grado.
60% SCLC expresan PAK2. Potencial diana terapeútica???
Revisión de las traslocaciones en NSCLC y métodos de
detección. Papel de la inmunohistoquímica.
Biomarcadores en citología.
Proposal for multicentre international standardization project
A multicenter, international standardization effort could address many of these questions and
help develop one “standardized” assay, for each of this family of drugs that comes into clinical
use as well as analyze additional immunotherapy-related predictive markers.
The IASLC Pathology Committee raises the prospect of trying to harmonize and standardize
testing for PD-L1 by IHC, at least at a technical level, but also, ideally, as a predictive marker, in
order to facilitate availability of this test and a promising treatment for patients with NSCLC.
Legend Figure 1 PDL-1 immunostaining performed using the E1LN3N clone anti-PD-L1 from
Cell Signaling Technology (Boston, USA) with standard detection techniques. A: Squamous cell
carcinoma showing a strong, uniform positive reaction in tumour cells. B: Despite being negative in tumor cells in the
centre of the image, there is a positive reaction in macrophages and other immune cells in the tumour stroma. C: Most
alveolar macrophages are positive for PD-L1. D: This adenocarcinoma is negative for PD-L1. It should be noted
that this IHC clone was not used for PD-L1 detection in any of the trials discussed in this
review.
ALK IQH como screening
 ALK IHQ negatico / no FISH
 ALK IHQ positivo / confirmación por FISH
Algunos ensayos clínicos aceptan ALK IHQ / VENTANA
+ neumocitos reactivos y CGMN
NRG1 (Neuregulin) “The newest kid on the block”
 Codifica una proteína de la familia EGF
 7-27% de Ac mucinosos invasivos
 Mujeres no fumadoras
 Ensayos clínicos en marcha frente posible diana (ERBB3 inhibitor)
Predictive biomarker testing in cytology ans small biopsy specimens
Nastasha Rekhman, MD, PhD
MSKCC, New York
•
Limitada celularidad
Retos y oportunidades
Infrautilización.
Preanalítica variable.
Optimización.
Obtención y manejo de
las muesras
Alto % de pacientes se diagnostican mediante estas muestras
Su uso adecuado permitiría mayor beneficio.
BRAF Mutations in NSCLC: Clinical Features and Outcome of a Clinical Series of Patients Diagnosed by Cytology
María D. Lozano M.D.1, Tania Labiano M.D. 1,José I Echeveste M.D.1 , Alfonso Gúrpide PhD 2, Nerea Gómez CLT1, . P. Preciado MD2, José Luis Solorzano M.D.1 , Hernán Quiceno M.D.1 ,
Mariam Maset CLT1, Salvador Martín-Algarra - Algarra MD2.
1. Department of Pathology, 2. Department of Medical Oncology. University Clinic of Navarra.
Pamplona, SPAIN
BACKGROUND
RESULTS
Presence of genetic alterations in various kinases is known as predictive markers in non-small cell lung carcinoma (NSCLC). The classification • BRAF wild type was found in 200 cases (97.6%), one was invalid due to paucity of DNA (0.5%) (Table 5). Four cases (2 %) harbored BRAF V600
schema is based on specific so-called driver mutations in frequencies exceeding 1%. BRAF is one of three members of the RAF kinase family.
mutation. DNA was obtained from Papanicoalou stained smears in two cases, one pleural fluid, and one cell block from a case for consultation. In
BRAF-mutations are known as malignant drivers in a number of cancers. While BRAF mutations in NSCLC have been described for several
these positive cases Cobas results were confirmed by direct sequencing. All harbored the V600E mutation (Figure 2). No concomitant EGFR and
years, the actual prevalence and clinical features of patients with NSCLC who harbor BRAF mutations are not well defined. Therapies against
KRAS mutations were detected. ALK rearrangements were not observed in the four BRAF mutated cases.
the specific V600-mutated BRAF-variant are developed and show promising results.
• All cases were stage IV adenocarcinoma, all patients were males and smokers. (Table 6)
We report a series of 205 consecutive NSCLC patients diagnosed by FNA on which BRAF mutational analysis was performed as a part of • They were enrolled in a clinical trial using selective inhibitors of mutant BRAF. All four patients had partial response and three persist up to date
routine molecular analysis.
in disease stabilization at 7, 14, and 16 moths.
PATIENTS AND METHODS
• Analysis of BRAF was introduced during last year as a part of routine molecular studies in NSCLC patients together with EGFR, KRAS, and ALK.
• We analyzed 205 cytological samples including 183 FNA, 8 pleural fluids, and 14 samples received for consultation. Sample procurement
details are summarize in Table 1.
• ROSE was performed in all cases to ensure a correct management of the samples. (Figure 1 )
• Analysis of BRAF was performed using Cobas 4800 Braf mutation test in 96 cases (46.8%), direct sequencing in 72 (35.1%), and
pyrosequencing in 28 (13.7%). Nine of these cases were analyzed in duplicated by Cobas and direct sequencing (Table 2).
• Clinicopathological characteristics are summarize in Table 3 and Table 4
Cytological samples
N
%
FNA-Bronchoscopy
54
26,3%
FNA-EBUS
38
18,5%
FNA-EUS
39
19%
FNA-CT
19
9,2%
FNA-Ultrasonography
17
8,2%
Direct superficial FNA
16
7,8%
Pleural Fluid
8
3,9%
Consultant cases*
14
6,8%
205
100%
*11 Stained Smears and 3 cell blocks
Figure 1: ROSE (Rapid On Site Evaluation)
Method for BRAF analysis
N
%
Cobas
96
46,8%
Direct Sequencing
72
35,1%
Pyrosequencing
28
9
4,4%
Total
205
100%
Table 4: Gender
Non-smoker
40
19,5%
Smoker ≤ 10 pack/year
14
6,8%
Smoker > 10 pack year
140
68,3%
Missing information
11
5,4%
Total
205
100%
#
%
Male
148
72,2%
Female
57
27,8%
Total
205
100%
200
4
2%
Invalid
1
97,6%
0,5%
Total
205
100%
V600E confirmed mutation
by direct sequencing
Positive
cases
Age
Gender
Histological subtype
Stage
Smoking
History
Status / Followup
1
69
Male
Adenocarcinoma
IV
60 pack/year
Dead*
2
72
Male
Adenocarcinoma
IV
40 pack/year
Alive
(7 months)
3
57
Male
Adenocarcinoma
IV
35 pack/year
Alive
(14 months)
4
59
Male
Adenocarcinoma
IV
60 pack/year
Alive
(16 months)
TTF-1
Smoking Status
Gender
WT
V600E mutation
Table 6: Clinicopathological Characteristics of four BRAF
mutated patients
13,7%
Cobas & Direct sequencing
Table 3: Smoking history
Table 5: BRAF status
BRAF
Table 2: BRAF mutation test
Table 1: Sample procurement
Total
Figure 2:Cytological findings in one of the cases harboring BRAF
V600E mutation
* Disseminated intravascular coagulation
CONCLUSIONS
To our knowledge this is the first clinicopathological study that includes BRAF in a routine comprehensive diagnostic panel of molecular drivers in
serial cytological samples of NSCLC. The frequency of BRAF mutation is 2%. All mutations were found in adenocarcinomas. Contrary to date
reported from most retrospective larger surgical series, mutations of BRAF occur in males and smokers.
Clinical trails show promising results in BRAF mutated patients. This study supports the value of cytological samples for comprehensive molecular
panels. Adequate management of samples is mandatory.
REFERENCES
1.Brustung OT, Khattak AM, Tromborg AK, Beigi M, Beiske K, Lund-Iversen M, Helland A. BRAF-mutations in non-small cell lung cáncer. Lung Cancer 2014;84:36-8
2.Li S, Li L, Zhu Y, Huang C, Qin Y, Liu H, Ren-Heidenreich L, Shi B, Ren H, Chu X, Kang J, Wang W, Xu J, Tang K, Yang H, Zheng Y, He J, Yu G, Liang N. Coexistence of EGFR with KRAS, or BRAF, or PIK3CA somatig mutations in lung cáncer: a comprehensive mutation profiling from 5125 Chinese cohorts. Br J Cancer 2014;110:2812-20
3. Villaruz LC, Socinski MA, Abberbock S, Berry LD, Jonhson BE, Kwiatkowski DJ, Iafrate AJ, Varella-Garcia M, Franklin WA, Camidge DR, Sequist LV, Haura EB, Ladanyi M, Kurland BF, Kugler K, Minna JD, Bunn PA, Kris MG. Clinicopathological features and outcomes of patients with lung adeocarcinomas harboring BRAF mutations in the Lung Cancer Mutation Consortium. Cancer 2015;121:448-56
4.Marchetti A, Felicione L, Malatesta S, Grazia Sciarrotta M, Guetti L, Viola P, Pullara C, Mucilli F, Buttitta F. Clinical features and outcome of patients with non-small-cell lung cáncer harboring BRAF mutations. J Clin Oncol 2011;29:3574-9
CONCLUSIONS
Table 5: BRAF status
BRAF
WT
V600E
mutation
Invalid
Total
200
97,6%
4
2%
1
205
0,5%
100%
To our knowledge this is the first clinicopathological study that includes BRAF in
a routine comprehensive diagnostic panel of molecular drivers in serial
cytological samples of NSCLC. The frequency of BRAF mutation is 2%. All
mutations were found in adenocarcinomas. BRAF Mmutations occur in males
and smokers.
Clinical trails show promising results in BRAF mutated patients. This study
supports the value of cytological samples for comprehensive molecular panels.
Adequate management of samples is mandatory.
Table 6: Clinicopathological Characteristics of four BRAF
mutated patients
Positive
cases
1
Age
Gender
69
Male
Histological
subtype
Adenocarcinoma
IV
Smoking
History
60 pack/year
2
72
Male
Adenocarcinoma
IV
40 pack/year
3
57
Male
Adenocarcinoma
IV
35 pack/year
4
59
Male
Adenocarcinoma
IV
60 pack/year
Stage
Status / Followup
Dead*
Alive
(7 months)
Alive
(14 months)
Alive
(16 months)
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Referencias