1111111111111111 IIIIII IIIII 111111111111111 11111 11111 111111111111111 IIIII 1111111111 11111111
`US 20120009121Al
`
`c19) United States
`c12) Patent Application Publication
`Pomper et al.
`
`c10) Pub. No.: US 2012/0009121 Al
`Jan. 12, 2012
`(43) Pub. Date:
`
`(54) PSMA-TARGETING COMPOUNDS AND USES
`THEREOF
`
`(51)
`
`(75)
`
`Inventors:
`
`Martin Pomper, Baltimore, MD
`(US); Ronnie Charles Mease,
`Fairfax, VA (US); Ray Sangeeta,
`Ellicott City, MD (US); Ying Chen,
`Timonium, MD (US)
`
`(73) Assignee:
`
`The Johns Hopkins University,
`Baltimore, MD (US)
`
`(21) Appl. No.:
`
`13/257,499
`
`(22) PCT Filed:
`
`Mar. 19, 2010
`
`(86) PCT No.:
`
`PCT /US2010/028020
`
`§ 371 (c)(l),
`(2), ( 4) Date:
`
`Sep.19,2011
`
`Related U.S. Application Data
`
`Publication Classification
`
`Int. Cl.
`A61K 51100
`A61K 49100
`C12N 5100
`C07D 311182
`C07D 403/06
`C07D 249104
`C07D 213153
`C07F 5102
`C07F 5100
`C07C 323122
`C07D 257102
`C07F 15100
`C12Q 1102
`C07D 311178
`
`(2006.01)
`(2006.01)
`(2006.01)
`(2006.01)
`(2006.01)
`(2006.01)
`(2006.01)
`(2006.01)
`(2006.01)
`(2006.01)
`(2006.01)
`(2006.01)
`(2006.01)
`(2006.01)
`
`(52) U.S. Cl. ........... 424/1.11; 435/29; 424/9.1; 435/325;
`549/388; 548/455; 549/382; 546/331; 546/13;
`556/1; 562/556; 540/474; 546/2; 548/255
`
`(60) Provisional application No. 61/161,485, filed on Mar.
`19, 2009, provisional application No. 61/161,484,
`filed on Mar. 19, 2009, provisional application No.
`61/248,067, filed on Oct. 2, 2009, provisional applica-
`tion No. 61/248,934, filed on Oct. 6, 2009.
`
`(57)
`
`ABSTRACT
`
`Prostate-specific membrane antigen (PSMA) targeting com-
`pounds are described. Uses of the compounds for imaging,
`therapy, cell sorting, and tumor mapping are also described.
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 1 of 27
`
`US 2012/0009121 Al
`
`30MIN
`
`3.5 h
`
`FIG.1
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 2 of 27
`
`US 2012/0009121 Al
`
`FIG. 2
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 3 of 27
`
`US 2012/0009121 Al
`
`.
`(.9
`LL
`
`

`

`> ....
`N ....
`....
`1,0
`0
`0
`--- 0
`N
`0 ....
`N
`rJJ
`c
`
`0 ....
`.i;...
`.....
`rJJ =(cid:173)
`0 ....
`N
`~
`? ....
`;'
`
`('D
`('D
`
`N
`
`-....J
`N
`
`0 =
`....
`~ .....
`
`(')
`
`0 = ""O = O" -....
`('D = ..... t ,= -....
`
`....
`~ .....
`
`(')
`
`~ .....
`""O
`
`N~OH
`
`H2
`
`o
`
`Ph~ O
`
`NH
`
`NH
`
`O Z'Ph
`
`Ht 0
`
`CN
`HO2c---..__ "
`
`COzH
`
`e
`
`H02C H ~ ~ H C02H
`.,_} 1 i.
`O ~
`TI NH ----HO2C _/ \....._/ '
`N N
`NJ
`rCOzH
`
`NHBoc
`
`-
`
`O, _,,l...__..,,........___..,.,.....,_
`O
`
`t
`o Id
`ir
`
`-
`
`NH
`
`Resin'
`
`~h
`... )=O
`
`Ph'-(NH
`
`O ✓ \....._/ "-COzH
`
`N N
`(N NJ
`)
`CO2H
`
`Ho2c---..__ "
`
`FIG 4
`
`•
`
`SR-V-100
`
`H
`
`H H
`
`COzH
`
`O
`
`{ij 11
`lnCl3 200 mM NaOAc, 60°G, 30 min
`(e} D?S-urea, TEA, DMF, rt, OVERNIGHT;
`(d) TFA, CHzClz, t, OVERNIGHT;
`(c} DOTA-tris(tert-butylester)-CO2H, HBTU, HOBT, DIEA, DMF;
`20% PERIDINE in DMF
`(b) 20% PERIDINE IN DMF; Fmoc-Phe-OH, HBTU, HOET, DIEA, DMF;
`(a) 20% PERIDINE IN DMF; Fmoc-Phe-OH, HBTU, HOH, DIEA, DMF;
`HO2C H N N H COzH
`.,_} Jl. i
`O ~
`
`1111n-[SR-V-100]
`
`H
`
`O
`
`_ll_,__.,,,-....__.,,,.......___.,,,-....__/
`O
`
`-
`
`-
`
`HO~N -
`
`)-/
`HN}=O Ph
`
`O =ZNH
`
`f
`
`Ph '-<NH
`
`0
`
`,--\....._/ "-CO2H
`
`r-C02H
`
`HO2C---..__ "
`
`(~ ~J .
`
`O=<
`
`HO~N~NH
`0 =(_NH
`
`-
`
`O
`
`••• )-/
`Ph
`HN
`~O
`
`0
`
`O
`0 =f' \....._/ '--io
`N/n1P'-
`(1111 ::,i~'I
`l)~:i)
`
`NH
`
`Ph
`
`O Q . 0 Q
`
`0
`
`c
`
`Resin1O~NHBoc
`
`Resin1C~NHBoc -
`
`:c-:.
`• HN
`Ph
`~o
`
`NH
`
`O
`
`NH2
`
`Ph
`
`b
`
`Ph
`
`O NH
`
`d...
`
`FmocHNY--
`
`ResinlO~NHBoc -
`a
`
`NHFmoc
`
`•
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 5 of 27
`
`US 2012/0009121 Al
`
`..c:
`
`00 -
`
`LO .
`(!)
`LL
`
`..c:
`......
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 6 of 27
`
`US 2012/0009121 Al
`
`■
`
`co
`(!)
`LL
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 7 of 27
`
`US 2012/0009121 Al
`
`FIG. 7
`
`

`

`0 ....
`QO
`.....
`rJJ =(cid:173)
`0 ....
`N
`~
`? ....
`;'
`
`('D
`('D
`
`N
`
`> ....
`N ....
`....
`N --0
`0 ....
`N
`rJJ
`c
`
`1,0
`0
`0
`
`-....J
`N
`
`.... 0 =
`.... 0 = ""O = O" -....
`('D = ..... t ,= -....
`
`~ .....
`
`(')
`
`~ .....
`
`(')
`
`~ .....
`""O
`
`FIG. 8
`
`INTEGRATED AREA FROM EMISSION SPECTRA IN THE REGION 780-900 nm USED
`FOR QUANTUM YIELD MEASUREMENTS, 775 nm EXCITATION WAVELENGTH USED,
`
`ODyc27=0.05 (). = 775 nm) Em SPECTRA AREA= 29509
`
`OD1cG=0.035 (A= 775 nm) Em SPECTRA AREA= 6276
`
`QUANTUM YIELD OF YC-27 3 IN WATER= 0.053
`
`REF Q.Y. (ICG IN WATER)= 0.016
`
`900
`
`800
`
`700
`
`600
`
`500
`
`WAVELENGTH (nm)
`
`,
`
`.,,-~
`,
`
`O.OOb.-------=--"-
`
`<C
`en co
`~ 0.02
`ci'3
`<.) z:
`w
`□ g_ 0.04
`
`0.0 ~
`=>
`0
`LU a:::
`U)
`<..:>
`w
`z:
`0.5 ~
`~
`t-
`w
`U) z:
`i:::
`1.0 ~
`
`~,,~
`
`\_
`'
`'
`
`I
`I
`I
`I
`I
`I
`I
`I
`I
`
`I
`I
`I
`I
`I
`I
`
`,,
`,
`
`I
`I
`I
`
`I
`I
`I
`I
`I
`,",
`
`ABSORPTION AND EMISSION SPECTRA OF YC-27 3 IN WATER
`
`

`

`0 ....
`.....
`rJJ =(cid:173)
`0 ....
`N
`~
`? ....
`;'
`
`1,0
`
`('D
`('D
`
`N
`
`> ....
`N ....
`....
`N --- 0
`0 ....
`N
`rJJ
`c
`
`1,0
`0
`0
`
`-....J
`N
`
`.... 0 =
`.... 0 = ""O = O" -....
`('D = ..... t ,= -....
`
`~ .....
`
`(')
`
`~ .....
`
`(')
`
`~ .....
`""O
`
`FIG. 9
`
`20
`
`18
`
`16
`
`14
`
`TIME (nsec)
`12
`10
`
`FLUORESCENCE LIFETIME = 443 psec
`
`2 4 6 8
`
`-40
`
`I
`
`I
`
`4
`
`0:::
`UJ
`ci5
`Cl
`:::,
`<C
`--'
`en
`
`20
`
`18
`
`16
`
`14
`
`12
`
`10
`
`0 2 4 6 8
`
`TIME (nsec)
`
`100
`
`101
`
`MONO-EXPONENTIAL MODEL.
`DECAY WAS ANALYZED WITH A
`THE FLUORESCENCE INTENSITY
`
`103[ I\\
`
`102
`
`\ ~
`
`INTENSITY -TIME DECAY OF YC-27 IN WATER
`
`104
`
`~
`I-
`UJ
`:z::
`en
`i:=
`;
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 10 of 27
`
`US 2012/0009121 Al
`
`- - - - - - - - - - - - - - - - - - - . - - 1 . -Q
`
`•
`
`<O
`'
`
`....
`
`00
`'
`
`C')
`1
`
`c::>
`...-t
`
`M'
`LL
`0
`:z
`0
`
`~ I-z
`C)
`w u ~
`.
`:z
`0
`&2.
`(9
`(.!)
`0
`LL
`....J
`
`c:::>
`c::>
`LC)
`cr,
`c-...
`
`c:::>
`c::>
`LC)
`er,
`...--
`
`c::>
`c::>
`' c:>
`r-
`...-
`
`A.LISN31NI 3~N38S3BOnl~
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 11 of 27
`
`US 2012/0009121 Al
`
`DORSAL
`
`VENTRAL
`
`PRE-INJECTION
`
`FIG. 11A
`
`• FIG. 11C
`• FIG. 11E
`• FIG. 11G
`• FIG. 111
`
`FIG. 118
`
`• FIG. 11 D
`• FIG. 11F
`• FIG. 11H
`• FIG. 11J
`
`FIG. 11K
`70.5 h Pl
`
`FIG. 11 L
`
`FIG. 11 M
`
`7.00E-1
`
`5.00E-1
`
`2.00E,1
`7.00E-2
`0.00E0
`
`FIG. 11N
`
`FIG. 110
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 12 of 27
`
`US 2012/0009121 Al
`
`FIG. 12A FIG. 12B
`
`FIG. 12A FIG. 12B
`
`. 9.79E-1
`
`· "I
`.".;'
`
`FIG. 12C FIG. 120 r
`
`FIG. 12C FIG. 120
`
`7.00E-1
`
`5.00E-1
`
`2.00E-1
`
`FIG. 12E FIG. 12F
`
`b:~~~i FIG. 12E FIG. 12F
`
`FIG. 12G • FIG. 12H
`
`FIG. 12G FIG. 12H
`
`FIG. 121
`
`FIG. 12J
`
`FIG. 121
`
`FIG. 12J
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 13 of 27
`
`US 2012/0009121 Al
`
`.....
`...c:
`00
`
`.
`C)
`LL
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 14 of 27
`
`US 2012/0009121 Al
`
`..c:
`C"?
`
`..c:
`Lt")
`C"'!
`
`~
`
`•
`
`(.9 -LL
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 15 of 27
`
`US 2012/0009121 Al
`
`..r::::.
`a...,
`
`..r::::.
`C"':)
`
`U")
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`(.9
`LL
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`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 16 of 27
`
`US 2012/0009121 Al
`
`..c
`Ln
`
`c.o
`~ .
`(!) -LL
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 17 of 27
`
`US 2012/0009121 Al
`
`:3 u..
`
`a.. a:
`
`.
`(9
`LL
`
`~~ :::iE c::
`:::$.. ~
`LQ (cid:173)
`_c:o
`cf:.<">
`:::iE >
`a.. u
`<'I >-
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 18 of 27
`
`US 2012/0009121 Al
`
`■
`
`(9
`LL
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 19 of 27
`
`US 2012/0009121 Al
`
`OMIN
`
`10MIN
`
`30MIN
`
`60MIN
`
`FIG. 19
`
`

`

`0 ....
`0
`N
`.....
`rJJ =(cid:173)
`0 ....
`N
`~
`? ....
`;'
`
`('D
`('D
`
`N
`
`> ....
`N ....
`....
`N --- 0
`0 ....
`N
`rJJ
`c
`
`1,0
`0
`0
`
`-....J
`N
`
`0 =
`....
`~ .....
`
`(')
`
`0 = ""O = O" -....
`('D = ..... t ,= -....
`
`....
`~ .....
`
`(')
`
`~ .....
`""O
`
`640
`
`620
`
`I
`
`I
`
`WAVELENGTH [nmj
`600
`560
`
`580
`
`I
`
`I
`
`540
`
`I
`
`o.o I I
`
`520
`
`--.... __
`
`' . -----,,
`
`.,..,..,,-
`
`-,_,
`
`I
`
`0.50
`
`AMPLITUDE
`NORMALIZED
`
`~
`,
`
`1.00,
`
`' ' ' ' ' '
`
`------.... ........ _,
`
`540 560 580 600 620 640 660 680
`---r--.-..---.....-....---,---,
`
`FIG. 20
`WAVELENGTH [nmj
`
`I
`I
`I
`
`,'
`
`0.50~
`
`I
`I
`I
`I
`I
`I
`I
`I
`I
`
`0.0 +-1
`
`580
`
`570
`
`WAVELENGTH [nm]
`560
`540
`
`550
`
`530
`
`520
`
`0
`
`0•3
`
`0.40
`0.50
`0.60
`0.70
`0.80
`
`1.00 AMPLITUDE
`NORMALIZED
`
`--------------------
`--UNMIXED #2
`-UNMIXED#1
`
`0.90 L-~------
`1.001',.
`AMPLITUDE
`NORMALIZED
`
`0.1uG ,.,....,,.Q Q._ 0.01uG
`10uG --...,oo ✓ 1uG
`
`GFP-QD-SET (TOGETHER)
`
`OD-SET (EX-500, EM-540)
`
`GFP-SET (EX-465, EM-520)
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 21 of 27
`
`US 2012/0009121 Al
`
`a:
`::z:
`::ii:
`<=> CV,
`
`a..
`:z:
`~
`c:>
`c.c
`
`UJ
`Cl
`u5
`:::>
`_J
`LL
`
`UJ
`Cl
`u5
`a..
`a..
`
`~
`
`N .
`(9
`LL
`
`_.I
`
`~ 0
`
`Cl
`
`g
`
`:z:
`UJ >
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 22 of 27
`
`US 2012/0009121 Al
`
`VENTRAL
`
`DORSAL
`
`PIP SIDE
`
`FLU SIDE
`
`180 MIN Pl
`
`CONTROL
`
`YCVlll-36
`(150 ug)
`
`FIG. 22
`
`

`

`> .....
`N .....
`.....
`1,0
`0
`0
`--- 0
`N
`0 .....
`N
`1J1
`
`c
`
`--..l
`N
`'""'I
`0
`N w
`.....
`ni
`1J1 =(cid:173) ni
`N
`0 .....
`N
`J'-1
`? .....
`~
`
`~ = ..... s· =
`~ = ::: ....
`~ = ..... s· =
`'E. ....
`t
`ni = .....
`~ = .....
`
`87.231
`11.54
`1.23
`0.00
`EVENTS % GATED
`
`9752
`1290
`137
`0
`
`I LR
`LL
`UR
`UL
`QUAD
`
`GATED EVENTS: 11179
`GATE:G1
`ACQUISITION DATE: 23-JUL-09
`FILE: LN CAP.006
`
`FIG. 23
`
`17.881
`82.03
`0.08
`0.02
`EVENTS % GATED
`
`1882
`8636
`8
`2
`
`I LR
`LL
`UR
`UL
`QUAD
`
`GATED EVENTS: 10528
`GATE:G1
`ACQUISITION DATE: 23-JUL-09
`FILE: PIP.004
`
`0.681
`99.19
`0.05
`0.08 •
`
`73
`10718
`5
`9
`
`EVENTS % GATED
`
`I LR
`LL
`UR
`UL
`QUAD
`
`GATED EVENTS: 10805
`GATE:G1
`ACQUISITION DATE: 23-JUL-09
`FILE: FLU.002
`
`104
`.
`
`103
`.
`
`FL 1-H
`102
`.
`
`101
`.
`
`100
`
`104
`
`103
`
`FL1-H
`102
`
`101
`
`100
`
`1
`
`100 =:;~ ..... " ..... : 100.
`
`101
`
`I
`
`LN CAP.006
`
`~ 102
`
`103
`
`104
`
`~ 102
`
`PIP.004
`
`103
`
`104
`
`FLU.002
`
`103
`
`104
`
`-------------1 LL
`
`.._.....
`
`101
`
`LL -----------1 LL
`~ 102
`
`101
`
`

`

`> ....
`N ....
`....
`N --0
`0 ....
`
`'-C
`0
`0
`
`N
`00
`('j
`
`-..J
`N
`0 -.
`,i;;..
`N
`~ .....
`00 =(cid:173)~
`0 ....
`N
`~
` ....
`
`N
`
`;(cid:173)?
`
`;::;· = ..... o· = "'d = S::
`
`0 =
`....
`= .....
`;::;·
`
`t 'E.
`~ = .....
`"'d = .....
`
`8694 · 80.171
`100.00
`10845
`REGION EVENTS % GATED
`GATED EVENTS: 10845
`GATE: G1
`ACQUISI ION DATE: 08-JAN-04
`FILE: PIPios.008
`
`IR2
`R1
`
`104
`
`103
`
`FL1-H
`102
`
`101
`
`FIG. 24
`
`)()100
`
`1(
`
`104
`
`103
`
`FL1-H
`102
`
`I
`
`101
`
`~ 102
`
`LL
`....I
`
`103
`
`PIP-pos.008
`
`PIP-neg.006
`
`101
`
`LL
`
`~ 102
`:::c
`
`103
`
`104
`
`3.88!
`100.00
`REGION EVENTS % GATED
`GATED EVENTS: 10540
`GATE: G1
`ACQUISITIO DATE: 08-JAN-04
`FILE: PIP-nei006
`
`409
`10540
`
`!R2
`R1
`
`AT PASSAGE 3:
`
`104
`
`pip36.002
`
`:x:: ~ 101 rn3
`
`102
`
`103
`
`104
`
`104
`
`103
`
`FL1-H
`102
`
`24.431
`100.00
`REGION EVENTS % GATED
`GATED EVENTS: 10624
`GATE: G1
`ACQUISITION DATE: 08-JAN-04
`FILE: FLU.004
`
`2595
`10624
`
`I R2
`R1
`
`PIP.004
`
`101
`
`~ 102
`:::c
`
`LL
`
`103
`
`104
`
`1.561
`100.00
`REGION EVENTS % GATED
`GATED EVENTS: 10480
`GATE: G1
`ACQUISITION DATE: 06-JAN-04
`FILE: FLU.002
`
`163
`10480
`
`I R2
`R1
`
`104
`11
`
`103
`
`1
`
`FL1-H
`102
`160
`101! .If
`100 __ !):1-~--;·. R2
`
`101
`
`FLU.002
`
`--' LL
`~ 102
`
`103
`
`104
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 25 of 27
`
`US 2012/0009121 Al
`
`SPECIMEN_001-Pip ZERO 1
`(x 1,000) 250
`200
`<!= 150
`~ 100
`50
`
`·'
`
`SPECIMEN_001-Pip ZERO 1
`105
`104
`<!=
`w
`a.. 103
`102
`
`50 100 150 200 250
`(x 1,000)
`
`FSC-A
`
`102 103 104 105
`FITC-A
`
`TUBE: Pip ZERO 1
`POPULATION
`ENTS
`1
`
`,
`
`P2
`P3
`
`SPECIMEN_001-Pip 1000-1
`
`(x 1,000) 250
`200
`<!= 150
`c...,
`~ 100
`50
`
`· 50 100 150 200 250
`(x 1,000)
`
`FSC-A
`
`TUBE: Pip 1000-1
`POPULATION
`ENTS
`1
`
`,
`
`P2
`P3
`
`TO
`FIG. 25B
`
`#EVENTS
`1,334,712
`991,055
`0
`55
`
`%PARENT %TOTAL
`100,0
`74.3
`0.0
`0.0
`
`74.3
`0.0
`0.0
`
`SPECIMEN_001-Pip 1000-1
`105
`104
`<!=
`w
`a.. 103
`102
`
`102 103 1o4 105
`FITC-A
`
`#EVENTS %PARENT %TOTAL
`100.0
`1,404,987
`74.3
`1,043,719
`0.0
`63
`0.0
`145
`
`74.3
`0.0
`0.0
`
`FIG. 25A
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 26 of 27
`
`US 2012/0009121 Al
`
`(x 1,000) 250
`200
`~ 150
`~ 100
`50
`
`SPECIMEN_001-Pip10-1
`
`SPECIMEN_001-Pip 10-1
`
`105
`
`UJ
`
`~ 104
`CL. 103
`102
`
`50 100 150 200 250
`(x 1,000)
`
`FSC-A
`
`102 103 104 105
`FITC-A
`
`TUBE: Pip 10-1
`POPULATION
`
`1iENTS
`
`1
`
`P2
`P3
`
`FROM
`FIG. 25A
`
`TO
`FIG. 25C
`
`#EVENTS
`1,389,523
`1,036.758
`1
`90
`
`%PARENT %TOTAL
`100.0
`74.6
`0.0
`0.0
`
`74.6
`0.0
`0.0
`
`SPECIMEN_001-Pip 10 K-3
`105
`104
`~
`UJ
`CL. 103
`102
`
`102 103 1o4 105
`FITC-A
`
`#EVENTS
`1,378,312
`1,039,977
`583
`775
`
`%PARENT %TOTAL
`100.0
`75.5
`0.0
`0.1
`
`75.5
`0.1
`0.1
`
`SPECIMEN_001-Pip 10 K-3
`
`(x 1,000) 250
`200
`~ 150
`~ 100
`50
`
`50 100 150 200 250
`(x 1,000)
`
`FSC-A
`
`TUBE: Pip 10 K-3
`POPULATION
`
`1iENTS
`
`1
`P2
`P3
`
`FIG. 258
`
`

`

`Patent Application Publication
`
`Jan. 12, 2012 Sheet 27 of 27
`
`US 2012/0009121 Al
`
`(x 1,000) 250
`200
`<f 150
`~ 100
`50
`
`(._)
`
`SPECIMEN_001-Pip 100-1
`
`SPECIMEN_001-Pip 100-1
`105
`
`UJ
`
`<f 1o4
`a.. 103
`102
`
`50 100 150 200 250
`(x 1,000)
`
`FSC-A
`
`102 103 104 105
`FITC-A
`
`TUBE: Pip 100-1
`POPULATION
`
`1iENTS
`
`P2
`P3
`
`FROM
`FIG. 258
`
`SPECIMEN_Q01-Pip 100-K3
`
`(x 1,000) 250
`200
`~ 150
`~ 100
`50
`
`#EVENTS
`1,416,263
`1,049,389
`9
`110
`
`%PARENT %TOTAL
`100.0
`74.1
`0.0
`0,0
`
`74.1
`0,0
`0.0
`
`SPECIMEN_001-Pip 100-K3
`105
`
`<C 1o4
`w
`a.. 103
`102
`
`TUBE: Pip 1 00-K3
`POPULATION
`
`1iENTS
`
`1
`P2
`P3
`
`50 100 150 200 250
`• (x 1,000)
`FSC-A
`
`102 103 104 105
`FITC-A
`
`#EVENTS
`1,376,175
`1,043,433
`4,974
`6,264
`
`%PARENT %TOTAL
`100.0
`75,8
`0.4
`0.5
`
`75.8
`0.5
`0.6
`
`FIG. 25C
`
`

`

`US 2012/0009121 Al
`
`Jan. 12, 2012
`
`1
`
`PSMA-TARGETING COMPOUNDS AND USES
`THEREOF
`
`CROSS-REFERENCE TO RELATED
`APPLICATION
`
`[0001] This application claims priority to U.S. Provisional
`Application Nos. 61/161,484 filed Mar. 19, 2009, 61/161,
`485, filed Mar. 19, 2009, 61/248,067, filed Oct. 2, 2009, and
`61/248,934, filed Oct. 6, 2009. The entire content of each
`Provisional Application is hereby incorporated by reference
`in their entirety.
`[0002] This invention was made using U.S. Government
`support under NIH grant NIH U24 CA92871. The govern(cid:173)
`ment has certain rights in this invention.
`
`BACKGROUND
`
`[0003]
`1. Field of the Invention
`[0004] The present invention relates to prostate specific
`membrane antigen (PSMA) binding compounds, chemical
`precursors of PSMA binding compounds and imaging meth(cid:173)
`ods of using the compounds.
`[0005] 2. Background
`[0006] Prostate cancer (PCa) is the most commonly diag(cid:173)
`nosed malignancy and the second leading cause of cancer(cid:173)
`related death in men in the United States (Cancer Facts &
`Figures; American Cancer Society: Atlanta, Ga., 2009). In
`2009, it is estimated that 192,000 men will be diagnosed with
`prostate cancer and 27,000 men will die of the disease. Only
`one half of tumors due to PCa are clinically localized at
`diagnosis and one half of those represent extracapsular
`spread. Localization of that spread as well as determination of
`the total body burden of PCa have important implications for
`therapy, particularly as new combination and focal therapies
`become available.
`[0007] The prostate-specific membrane antigen (PSMA),
`while expressed in prostate tumor epithelium, has a curious
`property in that it is expressed in the neovasculature of many
`solid tumors but not in that of prostate cancer (Chang et al.,
`Cancer Res., vol. 59, pp. 3192-3198, 1999; Chang et al., Clin.
`Cancer Res., vol. 5, pp. 2674-2681, 1999; Gong et al., Cancer
`Metastasis Rev., vol. 18, pp. 483-490, 1999; Chang et al.,
`Mo!. Ural., vol. 3, pp. 313-320, 1999; Baccala eta!., Urology,
`vol. 70, pp. 385-390, 2007; Chang et al., Urology, vol. 57, pp.
`801-805, 2001 Milowsky et al., J. Clin. Oneal., vol. 25, pp.
`540-547, 2007). Because of that property, an 111 In-labeled
`monoclonal antibody to an extracellular epitope of PSMA,
`111 In-J591, was capable of identifying renal, bladder, lung,
`breast, colorectal and pancreatic tumors in a Phase I clinical
`imaging study (Milowsky et al., J. Clin. Oneal., vol. 25, pp.
`540-547, 2007). That study validated 111 In-1591 as a vascular
`targeting agent in human subjects. Since then other reports
`have further studied PSMA expression in certain tumor types.
`Baccala et al. noted that clear cell renal cell carcinoma
`expresses significantly more PSMA in its neovasculature than
`does the papillary variety (Baccala et al., Urology, vol. 70, pp.
`385-390, 2007). Furthermore, angiomyolipoma, a benign
`renal lesion, did not express PSMA. As an enzyme with an
`extracellular active site, PSMA represents an excellent target
`for imaging and therapy directed toward solid tumor neovas(cid:173)
`culature in addition to prostate cancer itself. PSMA-based
`agents can report on the presence of this marker, which is
`increasingly recognized as an important prognostic determi(cid:173)
`nate inPCa (Murphy eta!., Urology, vol. 51, pp. 89-97, 1998).
`
`It is also the target for a variety of new PCa therapies (Gal sky
`etal.,JClin Oneal, vol. 26, pp. 2147-2154, 2008).
`[0008] ProstaScint™ is an 111 In-labeled monoclonal anti(cid:173)
`body against PSMA that is clinically available for imaging
`PCa. Radioimmunotherapy based on ProstaScint™ and
`radio labeled variations of this antibody are fraught with simi(cid:173)
`lar difficulties to the use of radio labeled antibodies for imag(cid:173)
`ing, including prolonged circulation times, poor target to
`nontarget tissue contrast, unpredictable biological effects and
`the occasional need for pre-targeting strategies, limiting the
`utility of these agents (Lange, P. H., Urology, vol. 57, pp.
`402-406, 2001; Haseman et al., Cancer Biother Radiopharm,
`vol. 15, pp. 131-140, 2000; Rosenthal et al., Tech Ural, vol. 7,
`pp. 27-37, 2001). Furthermore, antibodies may have less
`access to tumor than low molecular weight agents, which can
`be manipulated pharmacologically.
`[0009] The development of low molecular weight radio(cid:173)
`therapeutic agents is much different from developing radiop(cid:173)
`harmaceuticals for imaging in that longer tumor residence
`times can often be important for the former.
`[0010] Complete detection and eradication of primary
`tumor and metastatic foci are required to effect a cure in
`patients with cancer; however, current preoperative assess(cid:173)
`ment often misses small metastatic deposits. More sensitive
`imaging techniques than computed tomography, magnetic
`resonance imaging and even positron emission tomography
`(PET), which can be used easily in the operating suite, are
`required. An old technique, recently revisited because of
`improved optics and fluorescent dye chemistry, is intraopera(cid:173)
`tive photodiagnosis (PDD) (Toda, Keio J. Med., vol. 57, pp.
`155-161, 2008). Fluorescein dyes have been used intraopera(cid:173)
`tively to identify brain tumors and verify the clarity of tumor
`margins since 1948 (Toda, Keio J. Med., vol. 57, pp. 155-161,
`2008). A recent report describes its utility in identifying brain
`metastases (Okuda et al., Minim. Invasive Neurosurg., vol.
`50, pp. 382-384, 2007). A long history of the use of 5-ami(cid:173)
`nolevulinic acid (5-ALA) for brain tumor resection is also
`evident, and its use has been associated with improvement in
`progression-free survival (Stummer et al., Lancet Oneal., vol.
`7, pp. 392-401, 2006). PDD can be performed easily during
`surgery due to the lack of a need for complex imaging equip(cid:173)
`ment. All that is needed is a light-emitting diode to excite the
`fluorophore, which can be administered systemically or
`"painted" on the tissue directly. More recent incarnations of
`PDD have used quantum dots (Arndt-Jovin et al., IEEE Trans
`Nanobioscience, 2009), and more advanced dyes, such as
`indocyanine green (ICG) (Gotoh et al., J. Surg. Oneal., 2009),
`which emit in the near-infrared (NIR) region of the spectrum,
`enabling reasonable tissue penetration of emitted (and
`detected) light. Applications have included nontargeted
`approaches, such as preoperative evaluation of the vascular
`integrity of surgical flaps or identification of nodules ofhepa(cid:173)
`tocellular carcinoma (Matsui et al., Plast. Reconstr. Surg.,
`vol. 123, pp. 125e-127e, 2009). Targeted approaches are also
`emerging, such as use of a fluorophore-conjugated anti-CEA
`antibody to identify colon or pancreatic cancer (Kaushal et
`al., J. Gastrointest. Surg., vol. 12, pp. 1938-1950, 2008), or
`the use of NIR activatable probes that emit light only when
`cleaved by a tumor-associated protease (Sheth et al., Gynecol.
`Oneal., vol. 112, pp. 616-622, 2009).
`[0011] Recently, the application of 68Ga-labeled peptides
`has attracted considerable interest for cancer imaging
`because of the physical characteristics ofGa-68 (Reubi et al.,
`J Nucl Med, vol. 49, pp. 1735-1738, 2008). Ga-68 is available
`
`

`

`US 2012/0009121 Al
`
`Jan. 12, 2012
`
`2
`
`from an in-house 68Ge/ 68Ga generator (68Ge, t 112=270.8 day),
`which renders it independent of an onsite cyclotron. There(cid:173)
`fore, 68Ga-based PET agents possess significant commercial
`potential and serve as a convenient alternative to cyclotron(cid:173)
`based isotopes for positron emission tomography (PET), such
`as 18F or 1241. 68Ga has a high positron-emitting fraction (89%
`of its total decay). The maximum positron energy of 68Ga
`(max. energy=l.92 MeV, mean=0.89 MeV) is higher than
`that of 18F (max=0.63 MeV, mean=0.25 MeV). However, a
`study of spatial resolution using Monte Carlo analysis
`revealed that under the assumption of3 mm spatial resolution
`for most PET detectors, the full-width-at-half-maximum
`(FWHM) of 18F and 68Ga are indistinguishable in soft tissue
`(3.01 mm vs. 3.09 mm) (Sanchez-Crespo et al., Eur J Nucl
`Med Mal Imaging, vol. 31, pp. 44-51, 2004). That finding
`implies that with the standard spatial resolution of 5 to 7 mm
`for current clinical scanners, image quality using 68Ga-based
`radiotracers will likely be indistinguishable from that of 18F(cid:173)
`based agents, stimulating interest in the development of 68Ga(cid:173)
`labeled compounds for medical imaging (Sanchez-Crespo et
`al., EurJ Nucl Med Mal Imaging, vol. 31, pp. 44-51, 2004;
`Khanetal.,EurJSurgOncol, vol. 35,pp. 561-567, 2009; Fani
`et al., Contrast Media Mal Imaging, vol. 3, pp. 67-77, 2008).
`With a physical half-life of 68 min, 68Ga is also matched
`nicely to the pharmacokinetics of many peptides used for
`imaging. Few 68Ga-labeled, mechanism-based radiotracers
`for prostate cancer have been reported previously, and none
`for PSMA. Furthermore, 68Ga is introduced to biomolecules
`through macrocyclic chelators, which allows possible kit for(cid:173)
`mulation and wide availability of the corresponding imaging
`agents.
`
`SUMMARY OF THE INVENTION
`[0012] The present invention satisfies the long standing and
`unmet need for new imaging and therapeutic compounds for
`targeting prostate cancer and cancer angiogenesis. The
`present invention, in particular, provides therapeutic com(cid:173)
`pounds and imaging agents which differ from the prior art in
`modifications which were not previously known or sug(cid:173)
`gested. Furthermore, the invention provides imaging agents
`that offer better contrast between target tissues and non-target
`tissues. The invention also provides compounds with greater
`cellular retention and low molecular weight.
`[0013] Embodiments of the invention include compounds
`having the structure
`
`independently selected from hydrogen or a protecting group,
`a is 1, 2, 3, or 4, and R is each independently Hor C 1 -C4 alkyl.
`[0015] Variable r is O or 1. Tz is a triazole group selected
`from the group consisting of
`
`L'IB + (CH , ) ,+ m
`
`+ (CH , ) J : x •+
`
`X 1 is -NRC(O)-, -NRC(O)NR-, -NRC(S)NR-, or
`-NRC(O)O-; X2
`is ----C(O)NR-, -NRC(O)NR-,
`-NRC(S)NR-, or ---OC(O)NR-; R5 is H, CO2 H, or
`CO2R6
`, where R6 is a C 1-C 6 alkyl, C2 -C 12 aryl, or C4 -C 16
`alkylaryl; bis 1, 2, 3, or 4; and dis 1, 2, 3, or 4.
`[0016] Variable q is O or 1. Wis -NRC(O)-, -NRC(O)
`NR-, NRC(S)NR-, -NRC(O)O-, ---OC(O)NR-,
`---OC(O)-, ----C(O)NR-, or -C(O)O-; R2 and R3 are
`independently H, CO2H, or CO2R4
`, where R4 is a C1 -C6
`alkyl, C2 -C 12 aryl, or C4 -C 16 alkylaryl, wherein if one ofR2
`and R3 is CO2H or CO2 R2
`, then the other is H; n is 1, 2, 3, 4,
`5 or 6.
`
`[0014] wherein the subunits associated with elements p, q,
`r, ands may be in any order. Z is tetrazole or CO2 Q; each Q is
`
`[0017] Variable s is O or 1. Y is -C(O)-, -NRC(O)-,
`-NRC(S)-, -OC(O); and mis 1, 2, 3, 4, 5, or 6.
`
`

`

`US 2012/0009121 Al
`
`Jan. 12, 2012
`
`3
`
`[0018] Variable pis 0, 1, 2, or 3, and when pis 2 or 3, each
`R 1 may be the same or different. R 1 is H, C 1 -C6 alkyl, C2 -C 12
`aryl, or C4 -C 16 alkylaryl.
`[0019] G is a moiety selected from the group consisting of
`
`[0021] 2) when G is
`
`and r is 0, then q and s are both 0 or both 1;
`[0022] 3) when G is
`
`HO
`
`0
`
`HO
`
`then p is O and R 2 is H, and the structure optionally includes
`a chelated metal ion.
`[0023] 4) when G is
`
`where Ch is a metal chelating moiety, optionally including a
`chelated metal; FG is a fluorescent dye moiety which emits in
`the visible or near infrared spectrum; one of A and A' is Ch
`and the other is FG; V and V' are independently -C(O)-,
`-NRC(O)-, -NRC(S)-, or-OC(O)-; and g is 1, 2, 3,
`4, 5, or 6. The following conditions also apply:
`[0020]
`1) when G is
`
`andr is 0, then ifp is 0, then one ofR2 and R3 is CO2 R2
`the other is H; and
`[0024] 5) when g is
`
`, and
`
`and r is 0, then q and s are both 1;
`
`then r is 0.
`[0025] Embodiments include compounds having the struc(cid:173)
`ture
`
`

`

`US 2012/0009121 Al
`
`Jan. 12, 2012
`
`4
`
`wherein Z is tetrazole or CO2Q; each Q is independently
`selected from hydrogen or a protecting group, a is 1, 2, 3, or
`4, and R is each independently H orC 1 -C4 alkyl. Ch is a metal
`chelating moiety optionally including a chelated metal. W is
`-NRC(O)-, -NRC(O)NR-, NRC(S)NR-, -NRC(O)
`0-, -OC(O)NR-, ---OC(O)-, ----C(O)NR-, or----C(O)
`0-. Y is ----C(O)-, -NRC(O)-, -NRC(S)-, ---OC(O).
`Vis -C(O)-, -NRC(O)-, -NRC(S)-, or-OC(O)-.
`In exemplary embodiments mis 1, 2, 3, 4, 5, or 6; n is 1, 2, 3,
`4, 5 or 6; and pis 0, 1, 2, or 3, and when pis 2 or 3, each R 1
`may be the same or different. R 1 is H, C 1 -C6 alkyl, C2 -C 12
`aryl, or C4 -C 16 alkylaryl. R2 and R3 are independently H,
`CO2 H, or CO2R4
`, where R4 is a C 1 -C6 alkyl, C2 -C 12 aryl, or
`
`C4 -C 16 alkylaryl, wherein when one ofR2 and R3 is CO2 H or
`, the other is H, and when pis 0, one ofR2 and R3 is
`CO2R2
`CO2R4
`, and the other is H.
`[0026] Some embodiments further include a chelated
`metal. In some embodiments, the chelated metal is Tc, In, Ga,
`Y, Lu, Re, Cu, Ac, Bi, Pb, Sm, Sc, Co, Ho, Gd, Eu, Tb, or Dy.
`In some embodiments, the chelated metal an isotope, for
`example. In some embodiments, the isotope is Tc-94m,
`Tc-99m, In-111, Ga-67, Ga-68, Y-86, Y-90, Lu-177, Re-186,
`Re-188, Cu-64, Cu-67, Co-55, Co-57, Sc-47, Ac-225,
`Bi-213, Bi-212, Pb-212, Sm-153, Ho-166, or Dy-166.
`Embodiments include compounds having the structure
`
`0
`
`HO
`
`optionally including a chelated metal ion. Z is tetrazole or
`CO2Q; each Q is independently selected from hydrogen or a
`protecting group, and a is 1, 2, 3, or 4. R is each independently
`Hor C 1 -C4 alkyl.Wis -NRC(O)-, -NRC(O)NR-, NRC
`(S)NR-, -NRC(O)O-, -OC(O)NR-, ---OC(O)-,
`----C(O)NR-, or----C(O)O-. Y is ----C(O)-, -NRC(O)-,
`-NRC(S)-, -OC(O)-;
`[0027]
`In exemplary embodiments mis 1, 2, 3, 4, 5, or 6; n
`is 1, 2, 3, 4, 5 or 6; q is O or 1; ands is O or 1. R3 is H, CO2 H,
`or CO2R4
`, where R4 is a C 1 -C6 alkyl, C2-C 12 aryl, or C4 -C 16
`alkylaryl. Some embodiments further include a chelated
`metal ion. In some embodiments, the metal ion is Tc, Re, Cu,
`or Ga. In some embodiments, the metal ion is Tc-99m,
`Re-186, Re-188, Cu-64, or Ga-68. In some embodiments, the
`metal ion is Tc-99m.
`[0028] Embodiments include compounds having the struc(cid:173)
`ture
`
`

`

`US 2012/0009121 Al
`
`Jan. 12, 2012
`
`5
`
`where p, q, ands are in the order drawn, and q ands are either
`both O or both 1. Z is tetrazole or CO2Q; each Q is indepen(cid:173)
`dently selected from hydrogen or a protecting group, and a is
`1, 2, 3, or 4. FG is a fluorescent dye moiety which emits in the
`visible or near infrared spectrum. R is each independently H
`or C 1 -C4 alkyl. V is -C(O)- or -NRC(O)- or -NRC
`(S)-. Wis -NRC(O)-, -NRC(O)NR-, NRC(S)NR-,
`-NRC(O)O-, ---OC(O)NR-, -OC(O)-, -C(O)NR-,
`or-C(O)O-. Y is ----C(O)-, -NRC(O)-, -NRC(S)-,
`-OC(O). In exemplary embodiments mis 1, 2, 3, 4, 5, or 6;
`n is 1, 2, 3, 4, 5 or 6; pis 0, 1, 2, or 3, and when pis 2 or 3, each
`R 1 may be the same or different. R 1 is H, C 1 -C6 alkyl, C2 -C 12
`aryl, or C4 -C 16 alkylaryl. R2 and R3 are independently H,
`CO2 H, or CO2R2
`, where R2 is a C 1 -C6 alkyl, C2 -C 12 aryl, or
`C4 -C 16 alkylaryl, wherein when one ofR2 and R3 is CO2 H or
`CO2 R2
`, the other is H. In some embodiments, the fluorescent
`dye moiety emits in the near infrared spectrum.
`[0029] Embodiments include compounds having the struc(cid:173)
`ture
`
`wherein Z is tetrazole or CO2Q; each Q is independently
`selected from hydrogen or a protecting group, and a is 1, 2, 3,
`or 4. One of A and A' is Ch and the other is FG, where FG is
`a fluorescent dye moiety which emits in the visible or near
`infrared spectrum and Ch is metal chelating moiety option(cid:173)
`ally including a chelated metal. R is each independently H or
`C 1 -C4 alkyl. V or V' are independently ----C(O)-, -NRC
`(0)-, or -NRC(S)-. W is -NRC(O)-, -NRC(O)
`NR-, NRC(S)NR-, -NRC(O)O-, -OC(O)NR-,
`-OC(O)-, ----C(O)NR-, or -C(O)O-. Y is -C(O)-,
`-NRC(O)-, -NRC(S)-, ---OC(O).
`In exemplary
`
`embodiments mis 1, 2, 3, 4, 5, or 6; n is 1, 2, 3, 4, 5 or 6; and
`g is 1, 2, 3, 4, 5, or6; pis 0, 1, 2, or 3, and when pis 2 or 3, each
`R 1 may be the same or different. R 1 is H, C 1 -C6 alkyl, C2 -C 12
`aryl, or C4 -C 16 alkylaryl. R2 and R3 are independently H,
`CO2H, or CO2 R4
`, where R4 is a C 1 -C6 alkyl, C2-C 12 aryl, or
`C4 -C 16 alkylaryl, wherein when one ofR2 and R3 is CO2 H or
`CO2R 2
`, the other is H. In some embodiments, the fluorescent
`dye moiety emits in the near infrared spectrum. Some
`embodiments further include a chelated metal.
`[0030] Embodiments include compounds having the struc(cid:173)
`ture
`
`

`

`US 2012/0009121 Al
`
`Jan. 12, 2012
`
`6
`
`wherein subunits associated with p, q, r, and s may be in any
`order. Z is tetrazole or CO2 Q; each Q is independently
`selected from hydrogen or a protecting group, and a is 1, 2, 3,
`or 4. R is each independently Hor C 1 -C4 alkyl. In this exem(cid:173)
`plary embodiment r is 1. Tz is a triazole group having the
`structure
`
`X 1 is -NRC(O)-, -NRC(O)NR-, NRC(S)NR-, or
`-NRC(O)O-; X2 is ----C(O)NR-, -NRC(O)NR-, NRC
`(S)NR-, or -OC(O)NR-; R5 is H, CO2H, or CO2R6
`,
`where R6 is a C 1-C6 alkyl, C2 -C 12 aryl, orC4 -C 16 alkylaryl; b
`is 1, 2, 3, or4; and dis 1, 2, 3, or 4. In exemplary embodiments
`q is 0 or 1, Wis -NRC(O)-, -NRC(O)NR-, NRC(S)
`NR-,
`-NRC(O)O-,
`-OC(O)NR-,
`-OC(O)-,
`-C(O)NR-, or----C(O)O-;nis 1, 2, 3, 4, 5 or6; andR2 and
`, where R4 is a
`R3 are independently H, CO2H, or CO2 R4
`C 1 -C6 alkyl, C2 -C 12 aryl, or C4 -C 16 alkylaryl, wherein if one
`of R2 and R3 is CO2 H or CO2R2
`, then the other is H. In
`exemplary embodiments s is 0 or 1; Y is ----C(O)-, -NRC
`(0)-, -NRC(S)-, -OC(O); and mis 1, 2, 3, 4, 5, or 6. In
`exemplary embodiments pis 0, 1, 2, or 3, and when pis 2 or
`3, each R 1 may be the same or different; and R 1 is H, C 1 -C6
`alkyl, C2 -C 12 aryl, orC4 -C 16 alkylaryl. G 1 is a moiety selected
`from the group consisting of
`
`-continued
`A'-V'
`
`R
`r-(CH2)g
`
`-......_-1yNH i
`N--._j
`
`A-V
`
`/ " - -
`
`0
`
`where Ch is a metal chelating moiety, optionally including a
`chelated metal; FG is a fluorescent dye moiety which emits in
`the visible or near infrared spectrum; one of A and A' is Ch
`and the other is FG; V and V' are each independently
`----C(O)-, -NRC(O)-, -NRC(S)-, or -OC(O)-; and
`g is 1, 2, 3, 4, 5, or 6. In some embodiments, the fluorescent
`dye moiety emits in the near infrared spectrum. Some
`embodiments include a chelated metal.
`[0031] Embodiments of the invention include methods of
`imaging one or more cells, organs or tissues by exposing the
`cell to or administering to a organism an effective amount of
`a compound discussed above, where the compound includes
`a fluorescent dye moiety, or a metal isotope suitable for imag(cid:173)
`ing.
`[0032] Embodiments of the invention include methods of
`treating a tumor comprising administering a therapeutically
`effective amou