Positive Bone Marrow Biopsy is Associated with a Decreased Disease-Free Survival in Patients with Operable Breast Cancer

Positive Bone Marrow Biopsy is Associated with a Decreased Disease-Free Survival in Patients with Operable Breast Cancer L Fortunato (*), A Mascaro (*), A Baldi ( ), M Farina (*), MAmini(**) ), C Vitelli (*) Department of Surgery (*), and Pathology (**) San Giovanni-Addolorata Hospital, Rome, Department of Biochemistry ( ), Second University of Naples ITALY SSO Annual Symposium Phoenix, AZ - March 7th, 2009 1

Background Staging of breast cancer patients is essential, but our ability to predict relapse is imprecise 30% of node negative patients will relapse 40% of node positive patients will survive long term Further prognostic factors are urgently needed SSO 2009 2

Review To date, at least 28 studies involving 6202 patients 18 of them have shown a correlation with DFS/OS Often, BM independent prognostic factor Lack of standardization and wide range of positivity Only few studies have employed RT-PCR SSO 2009 3

Aim of Study Bone Marrow: Since 2000 Peripheral blood: Since 2002 RT-PCR: CEA and Mammaglobin Primary end point: Secondary end point: disease-free survival overall survival correlation with Microarray SSO 2009 4

Methods Test was considered positive if any RT-PCR was positive All patients signed an informed consent Results remained unavailable to both patients and clinicians SSO 2009 5

Results 274 patients were enrolled BM biopsy was unsuccessful in 9 patients RT-PRC technically not feasible in 18 patients 247 patients t available for results Only one patient was lost to follow-up (0.4%) SSO 2009 6

Results RT-PCR Test was positive in 110/247 patients (45%) At a median F/U of 60 months 31 events (13%) have occurred 13 deaths due to disease 15 systemic recurrences 3 regional recurrences SSO 2009 7

Positive Test and Tumor Characteristics N BM + % p Age 049 0.49 < 50 50 24 48 > 50 196 86 44 Diameter 0.87 < 1 cm 64 27 42 1-2 cm 116 51 44 > 2 cm 62 29 47 Histology 0.93 Ductal 212 94 44 Non Ductal 34 16 47 Nodal Status 0.004 N0 139 51 37 N+ 107 59 55 Hormonal status 0.66 Positive 202 87 43 Negative 40 20 50 C-Erb B2 0.39 Positive 42 21 50 Negative 196 84 43 Molecular Type 0.31 Luminal like 187 87 46 Non Luminal Like 34 19 56 SSO 2009 8

Positive Test and Stage 80 70 60 50 40 30 20 STAGE I STAGE II STAGE III P = 0.006 SSO 2009 9

Disease free survival Univariate i analysis Hazard Ratio Lower 95% CI Upper 95% CI P Age 1 0.96 1.03 1 Menopausal status 08 0.8 027 0.27 233 2.33 069 0.69 Histology 0.51 0.12 2.19 0.36 Tumor diameter 2.93 0.94 9.08 0.06 Nodal status 2.24 1.1 4.8 0.03 Grade 4.53 1.81 11.4 0.001 Hormonal status 0.3 0.16 0.82 0.01 C-Erb B2 2.68 1.18 6.07 0.01 RT-PCR Test 4 1.7 9.4 0.001 SSO 2009 10

Disease free survival Univariate i analysis Hazard Ratio Lower 95% CI Upper 95% CI P Age 1 0.96 1.03 1 Menopausal status 08 0.8 027 0.27 233 2.33 069 0.69 Histology 0.51 0.12 2.19 0.36 Tumor diameter 2.93 0.94 9.08 0.06 Nodal status 2.24 1.1 4.8 0.03 Grade 4.53 1.81 11.4 0.001 Hormonal status 0.3 0.16 0.82 0.01 C-Erb B2 2.68 1.18 6.07 0.01 RT-PCR Test 4 1.7 9.4 0.001 SSO 2009 11

Disease free survival Multivariate Analysis Hazard Ratio Lower 95% CI Upper 95% CI P RT-PCR Test 354 3.54 103 1.03 12.12 004 0.04 Nodal status 1.7 0.38 7.58 0.49 Interaction Test / 0.99 0.17 5.73 0.99 Nodal status SSO 2009 12

Disease free survival According to RT-PCR test and Nodal status P< 0.001 P= 0.03 13

Bone Marrow and Peripheral blood Disease-free survival P = 0.0003 P = 0.78 14

CEA and Mammaglobin Disease free survival P < 0.001 P = 0.11 15

5 year Disease-free survival 100 95 90 85 80 75 70 65 60 55 50 N- / BM - N- / BM + N+ / BM - N+ / BM+ P= 0.005 SSO 2009 16

Re-staging? N- /BM- N- /BM+ N+ /BM- N+ /BM+ SSO 2009 17

Conclusions Positive RT-PCR of the BM for Mammaglobin is associated with a four-fold increased risk of death or relapse RT-PCR of the PB is not associated with DFS Patients N-/BM- are almost certainly cured Patients N+/BM+ need more aggressive strategies SSO 2009 18

19

Review Author Year N Method Compart Positive (%) DFS OS Multivariate Porro 1988 159 IHC Bone Marrow 16 - - N/A Harbeck 1994 100 IHC Bone Marrow 38 + + + Diel 1996 737 IHC Bone Marrow 43 + + + Landys 1998 128 IHC Bone Marrow 19 + + N/A Untch 1999 581 IHC Bone Marrow 28 - - N/A Mansi 1999 350 IHC Bone Marrow 25 + + + Braun 2000 552 IHC Bone Marrow 36 + + + Gerber 2001 507 IHC Bone Marrow 32 + + + Gebauer 2001 393 IHC Bone Marrow 42 + + + Solomayer 2001 727 IHC Bone Marrow + + + Wiedswang 2003 817 IHC Bone Marrow 13 + + + Wong 2003 371 IHC Bone Marrow 33 + - - Pierga 2008 621 IHC Bone Marrow 15 + + + Ooka 2001 111 RT-PCR BoneMarrow 30 + - + Stathopoulou 2002 148 RT-PCR Peripheral 30 + + + Farmen 2008 195 RT-PCR Bone Marrow 12 + - + Apostolaki 2008 216 RT-PCR Peripheral 24 + + + 20

1 2 3 4 5 C+ BM- BM- BM+ BM+ 21

Comparison of the power to identify tumor cells between tests 1000000 900000 800000 700000 600000 500000 400000 300000 200000 100000 0 H/H IHC RT-PCR 22

Eventi sistemici e Decessi 25 20 15 / 112 15 Eventi sistemici 7 / 112 Decessi 10 2 / 99 2 / 99 5 0 MIDOLLO POSITIVO MIDOLLO NEGATIVO Eventi: P= 0.009 Sopravvivenza p= 0.29 23

RT-PCR Genomic DNA Primer 1 Primer 1 Primer 3 Exon 1 Exon 2 Exon 3 Primer 4 Primer 2 cdna Primer 1 Primer 3 Exon 1 Exon 2 Exon 3 Primer 4 Primer 2 24

25

Sensitivity of Molecular testing by RT-PCR 80 70 60 50 40 30 20 10 0 MAMM PIP PSE CEA CK19 MUC1 Gillanders WE, et al: Ann Surg 2004; 239: 828-840 26

Comparazione tra la capacità di identificazione delle cellule tumorali 1000000 900000 800000 700000 600000 500000 400000 300000 200000 100000 0 E/E IHC RT-PCR 27

Tipo di test effettuato MIDOLLO + PERIFERICO SOLO PERIFERICO SOLO MIDOLLO Pazienti sono state considerate negative in assenza di positività in tutti i test (Midollo/periferico CEA/Mammolobina) 28