THETWO TORTOITUUSN 20180252720A1ULLUM HOLATIN ( 19) United States (12 ) Patent Application Publication ( 10) Pub . No. : US 2018 / 0252720 A1 DIEU -NOSJEAN et al. (43 ) Pub. Date : Sep . 6 , 2018 (54 ) METHODS FOR PREDICTING THE Publication Classification SURVIVAL TIME OF PATIENTS SUFFERING (51 ) Int . Ci. FROM A LUNG CANCER GOIN 33 /574 ( 2006 .01 ) (71 ) Applicants : INSERM (INSTITUT NATIONAL ( 52 ) U . S . CI. DE LA SANTE ET DE LA CPC .. GOIN 33 /57423 (2013 . 01) ; GOIN 2800/ 52 RECHERCHE MEDICALE ) , Paris ( 2013 . 01 ) (FR ) ; UNIVERSITE PARIS DESCARTES , Paris ( FR ) ; SORBONNE UNIVERSITE , Paris (57 ) ABSTRACT (FR ) ; UNIVERSITE PARIS DIDEROT - PARIS 7 , Paris ( FR ) ; The present invention relates to methods for predicting the ASSISTANCE survival time of patients suffering from a lung cancer . In PUBLIQUE -HOPITAUX DE PARIS particular , the present invention relates to a method for (ADHP ) , Paris (FR ) predicting the survival time of a subject suffering from a lung cancer comprising the steps of i) quantifying the ( 72 ) Inventors: Marie - Caroline DIEU - NOSJEAN , density of regulatory T ( Treg ) cells in a tumor tissue sample Paris (FR ) ; Wolf Herdman obtained from the subject, ii ) quantifying the density of one FRIDMAN , Paris ( FR ) ; Catherine further population of immune cells selected from the group SAUTES - FRIDMAN , Paris ( FR ) ; consisting of TLS -mature DC or TLS - B cells or Tconv cells , Priyanka DEVI, Paris (FR ) CD8 + T cells or CD8 + Granzyme - B + T cells in said tumor tissue sample , iii ) comparing the densities quantified at steps (21 ) Appl. No. : 15/ 754 , 640 i ) and ii ) with their corresponding predetermined reference values and iv ) concluding that the subject will have a short ( 22 ) PCT Filed : Aug . 26 , 2016 survival time when the density of Treg cells is higher than its corresponding predetermined reference value and the ( 86 ) PCT No .: PCT/ EP2016 /070158 density of the further population of immune cells is lower $ 371 ( c ) ( 1 ) , than its corresponding predetermined reference value or ( 2 ) Date : concluding that the subject will have a long survival time Feb . 23 , 2018 when the density of Treg cells is lower than its correspond (30 ) Foreign Application Priority Data ing predetermined reference value and the density of the further population of immune cells is higher than its corre Aug . 27 , 2015 ( EP ) .. .. .. 15306318 .5 sponding predetermined reference value . TTT %OverallSurvival to Tregs lo ( n = 53 ) Tregs Hi( n = 190 ) P = 0 .0049 0 20 40 60 80 100 120 140 Time in months OverallSurvival. TLS Tregs to ( n = 58) TLS Tregs Hi ( n = 42 ) P = 0 . 0245 TTTTT 0 20 40 60 80 100 120 140 Tima In months Patent Application Publication Sep . 6 , 2018 Sheet 1 of 17 US 2018 / 0252720 A1 - o Urvi NOUBLIA . LOOPINI TLMJDUJJDLOOLID III . Ñ L Tregs lo ( n = 53 ) Tregs Hi (n = 190 ) P = 0 .0049 0 20 40 60 80 100 120 140 Time in months · . * * * D * * * * ' D' SO %OverallSurvival . DEI wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww ' TLS Tregs lo ( n = 58 ) IDEO TLS Tregs Hi ( n = 42 ) P = 0 . 0245 0 20 40 60 80 100 120 140 Time in months Figure 1A - B Patent Application Publication Sep . 6 , 2018 Sheet 2 of 17 US 2018 / 0252720 A1 0 ODBO OverallSurvival — — ??????????????? ??????????????? ??????????. wwwwwwwwwww w wwwwwwwwwwwwww IWWWWWWWWWWWW DC Hi ( n = 138) DC lo ( n = 105 ) P = 0 . 0002 O TT 0 20 40 60 80 100 120 140 Time in months WWWWWWWWW o %OverallSurvival www ** ** * * ** * * ** * * * * * * * * ** ** * * * * * * * -. - to - HIDC / la Tregs ( n = 26 ) HIDC / Hi Tregs ( n = 11Z ) LO DC / LO Tregs ( n = 27 ) LODC / Hi Tregs ( n = 78 ) P < 0 .0001 0 20 40: 60 80 100 120 140 Time in months Figures 1C - D Patent Application Publication Sep . 6 , 2018 Sheet 3 of 17 US 2018 / 0252720 A1 P . e . www . .LLLLLLL . * * * * * * * * * ?????????????? ” ??? . OverallSurvival . wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww . B cell Hi ( n = 172 ) . B cell lo ( n = 71 ) . P = 0 . 0020 0 20 40 60 80 100 120 140 Time in months IIIIIIIIII IIIIIII IIIIIIIIIIII IBIIIIIIII %OverallSurvival HiB cells / lo Tregs ( n = 31) HiB cells / Hi Tregs ( n = 141 ) LOB cells / Lo Tregs ( n = 22 ) LOB cells / Hi Tregs ( n = 49 ) P < 0 .0001 T 0 20 40 60 80 100 120 140 Time in months Figure 1 E - F Patent Application Publication Sep . 6 , 2018 Sheet 4 of 17 US 2018 / 0252720 A1 64 IIIIIIII E IEEELD T . DIRIBEROI BÖLIBIIIIIIIII 13 OverallSurvival ??????????? À CD8 Hi ( n = 178 ) CD8 lo ( n = 63 ) P = 0 . 0027 TTTTTTTTTTT 0 20 40 60 80 100 120 140 0 IIIIIIIIIIIIIIIII IIIIIII %OverallSurvival to Hi CD8 /lo Tregs ( n = 37 ) Hi CD8 / Hi Tregs ( n = 141 ) Lo CD8 / Lo Tregs ( n = 16 ) o Lo CD8 / Hi Tregs ( n = 47 ) P = 0 . 0002 20 40 60 80 100 120 140 Time in months Figures 1G - H Patent Application Publication Sep . 6 , 2018 Sheet 5 of 17 US 2018 / 0252720 A1 Optimal cut- off value 1 . 12] . + + + + . T . 0.1 . + . + . + 0 .05 . + . + anjead + + . + . + + + + + + . + 100 . + . + . + . + . + . + . + . + . 898 . 7939 cells /mm2 . 0.001 . IIIIIIIIIIIIIIIIII I wo OZ+L 2300- . 008 2900 Density of CD3 + T cells ( cells /mm2 ) Optimal cut - off value - tait elen k - 01 - 0 .05 * 0.01 ILETILI+T11T1P1II1III11ETTIITTI ** p-value 11111111111 - 04000118- - -. 1e-05 21. 99746 cells /mm ? UTWELL L8L . Density of FoxP3+ Treg cells (cells / mm2 ) Figures 2A - B Patent Application Publication Sep . 6 , 2018 Sheet 6 of 17 US 2018 / 0252720 A1 Optimal cut- off value Ww.V RARLASERRISSURMARN A SS. 0.1 3 W . AWI Www Way Town where s ubteran 0 . 05 . MITTTTTTTT . W : 0.01 SHIRASHIFRA 001.0 L11MLIMILO .L 16-04 17 . 94117 cells /mm2 Density of the CD3 + T cells : Tregs ratio ( cells / mm2) Optimal cut- off value www ace. HO . * * * : . MIX . : : s it * * . t t . s . 0 . 05 . * anjeu-d 100 * * : n 1000 . i : 10-01 12 . 41506 cellselefon /mm2 LLLLLLLLLLLLL Density of the CD8 + T cells : Tregs ratio ( cells /mm2 ) Figures 2C - D Patent Application Publication Sep . 6 , 2018 Sheet 7 of 17 US 2018 / 0252720 A1 Optimal cut- off value L AMINA . 1SASHAUNAS 0.1 *withthe 11 : 01 TI . W . W e will be ablew ith Wale Wut wa linew ith p-value . .the ,. 0.001 . 0 . 06297162 cells /mm2 1e-04 . LLLLLLLLLLLLLLL Density of mDC cells : Treg cells (cells /mm2 ) horen en meer Optimal cut- off - value MITT MMWWWW WWW content contre este comentariosm sobre esteen esemegen : 0 .05 # D-value 0.01 0.001 LLL #DOT . LAALALLAAMAALAALUSVAATSULANTAIWAN 041e- 3.47.. 127. 0348 cells /mm2 Polda 1100- -1400 DensityDensite of the tres Tregsegs in horis fcellarells /mm2 m2) Figures 2E - F Patent Application Publication Sep . 6 , 2018 Sheet 8 of 17 US 2018 / 0252720 A1 . .. .. Piw ow . .wwwwwwwwwwwwwwwww %OverallSurvival Wwwx . www .w . w .w . www .w . www . www . www . www .wwwwwwwwwwwwwwwwww wwwww E USTATISTIMITS cD3 Hi ( n = 87 ) CD3 lo ( n = 156 ) P = 0 . 0073 p por r op 0 20 40 60 80 100 120 140 Time in months SIIRRITOTT . WSOUNDSYSSESSOUSCOUSSSSSSSSSSSSSSSSSSSS %OverallSurvival AKASSAR W . .. M SHAGANDA . CD3: FoxP3 ratio lo (n = 152) P = 0 . 0008 W 0 20 40 60 80 100 120 140 Time in months Figure 3 A - B Patent Application Publication Sep . 6 , 2018 Sheet 9 of 17 US 2018 / 0252720 A1 JUD . O OOOOOO . .. ) %QuerallSurvival . : . DC :FoxP3 ratio Hi (n = 66 ) DC : FoxP3 ratio lo ( n = 177 ) P = 0 . 0003 ?????. ???? ? ? ??? ???? ???? ?????? ?????? . ??? ?? ????? . ? ?? ????????????? ? ?????? ? o 20 40 60 80 100 120 140 Time in months . 0 . 0 . ILLIUS JOO . %OverallSurvival . .. ** * . CD8 :FoxP3 ratio Hi (n = 77 ) . CD8: FoxP3 ratio lo (n = 164 ) P = 0 .0005 0 0 20 40 60 80 100 120 140 Time in months Figure 3C - D Patent Application Publication Sep . 6 , 2018 Sheet 10 of 17 US 2018/ 0252720 A1 ? ? . - . .. - . - - - - - - -- 2 Tregs- Low ( n = 101 ) - 1 - Tregs Hi ( n = 237) all - All patients ( n = 338) P = 0 . 004 ?? : : : : ! (%)so ? ? ? ? ? ? ?? . r ? ? ? ? ??8?9???? ?? ? ? ??? ? 20 40 0 " " " 80" 1 100 120 Time ( months) B 2 - TLS- Tregs Low ( n = 58 ) -L. 1 - TLS - Tregs Hi ( n = 42) all - - All patients( n = 100) P = 0. 024 (%)so ??8090??? - ? ? ? LEl . III ? ? ?? ????? ?? ?? ? ? ? ? ? ? 20 40 60 80 100 120 Time ( months) Figure 4 A - B Patent Application Publication Sep . 6 , 2018 Sheet 11 of 17 US 2018 / 0252720 A1 o 1.0 - CD3 Hi ( n = 94) ] 2 – CD3 Low ( n = 244 ) 0.8 NH all - All Patients (n = 338) P = 0 .002 0.6 modo RA (%)SO Mwstgraukti?tinggit 1 .44 TEHT IH ! em * * *MW Hort 1 0.4 thangrant that 1 TEN111 ! + 2 0.2 HI 0. TTTT20 40 60 80 100 120 Time( months ) 1.0 TTTTTT 11111111111111111 0.8 mwater tanker off # 1 0.6 (%)50 MTTTILL HHHHHHHHHHIHI + H + + + + LI: 40. Cp3St Hl / Tregs Low ( n = 19 ) ht - H3all CD3 Hi/ Trags Hi (n = 75 ) 2 20. CD3 Low / Trags Low ( n = 82 ) 3 2 with + 4 IT CD3 Low / Trags HI ( n = 182 ) 4 All Pationts ( n = 338 ) P < 0 . 001 0. 0 20 40 60 80 100 120 Time (months ) Figure 4 C - D Patent Application Publication Sep . 6 , 2018 Sheet 12 of 17 US 2018 / 0252720 A1 E 1.0 1 - CD8 Hi ( n = 265) 2 -- CD8 Low (n = 70 ) 0.8 all - AllPatients ( n = 335 ) P < 0 . 001 0.6 OS(%) HILTILL 4111111 IT 0.4 * 1 all 0.2 barrisThe truthtorney fathe + # 2 0. 20 40 60 80 100 120 Time (months ! 1.0 1 B6Hit Fregs to th - bot F CD8 Hi/ Tregs Hi ( n = 197 ) CD8 Low / Tregs Low ( n = 32 ) 0.8 CDB Low / Tregs Hi ( n = 38 ) All Patients ( n = 33B ) 0.6 05(%) memoration that ITH + THrrr # 1 0.4 to benght 0.2 P << 0 . 001 0. TT 20 40 60 80 100 120 Time( months ) Figure 4 E - F Patent Application Publication Sep . 6 , 2018 Sheet 13 of 17 US 2018 /0252720 A1 G 1.0 1 - DC Hi ( n = 187) 2 - DC Low (n = 151) 0.8 all - All Patients ( n = 338 ) P < 0 .