Extended Time-lapse Intravital Imaging of Real-time Multicellular Dynamics in the Tumor Microenvironment

In the tumor microenvironment, host stromal cells interact with tumor cells to promote tumor progression, angiogenesis, tumor cell dissemination and ...

Albert Einstein College of Medicine

The B cell, a major component of humoral immunity, is a sensitive target for the immunotoxic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), possibly by rendering cells less responsive to antigenic or mitogenic stimulation. Potential mechanisms of TCDD action on B cells were examined in murine B cell lymphoma cells (CH12.LX) treated with 3 nM TCDD or dimethyl sulfoxide vehicle using sequence-verified cDNA microarrays. One transcript that was significantly induced by TCDD was suppressor of cytokine signaling 2 (Socs2). Changes in Socs2 mRNA levels paralleled that of Cyp1a1 with a maximal 3-fold induction observed at 4 h, as determined by quantitative real-time polymerase chain reaction. Socs2 induction seems B cell-specific, because no induction was observed in TCDD-responsive mouse hepatoma cells or human breast cancer cells. TCDD-mediated induction of Socs2 mRNA was dose-dependent and exhibited the characteristic structure-activity relationships observed for the aryl hydrocarbon receptor (AhR) ligands 3,3',4,4',5-pentachlorobiphenyl (PCB-126), indolo[3,2-b]-carbazole, and beta-naphthoflavone. Experiments with cycloheximide and AhR-deficient B cells indicated that Socs2 mRNA induction is a primary effect that is AhR-dependent. Western blot analysis confirmed that Socs2 and Cyp1a1 protein levels were also induced in CH12.LX cells. Promoter analysis revealed the presence of four dioxin-response elements within 1000 base pairs upstream of the Socs2 transcriptional start site, and a reporter gene regulated by the Socs2 promoter was inducible by TCDD. Promoter activity was also dependent on a functional AhR signaling pathway. These results indicate that Socs2 is a primary TCDD-inducible gene that may represent a novel mechanism by which TCDD elicits its immunosuppressive effects.

2,3,7,8-Tetrachlorodibenzo-p-dioxin induces suppressor of cytokine signaling 2 in murine B cells.

A transition metal complex targeted for the inhibition of a subset of zinc finger transcription factors has been synthesized and tested in Xenopus laevis. A Co(III) Schiff base complex modified with a 17-bp DNA sequence is designed to selectively inhibit Snail family transcription factors. The oligonucleotide-conjugated Co(III) complex prevents Slug, Snail, and Sip1 from binding their DNA targets whereas other transcription factors are still able to interact with their target DNA. The attachment of the oligonucleotide to the Co(III) complex increases specificity 150-fold over the unconjugated complex. Studies demonstrate that neither the oligo, or the Co(III) Schiff base complex alone, are sufficient for inactivation of Slug at concentrations that the conjugated complex mediates inhibition. Slug, Snail, and Sip1 have been implicated in the regulation of epithelial-to-mesenchymal transition in development and cancer. A complex targeted to inactivate their transcriptional activity could prove valuable as an experimental tool and a cancer therapeutic.

Targeted inhibition of Snail family zinc finger transcription factors by oligonucleotide-Co(III) Schiff base conjugate.

Eine chirale Porphyrazine (Pz), H(2)[pz(trans-A(2)B(2))] (247), dass Exponate bevorzugte die in-vivo Akkumulation in den Zellen von Tumoren erarbeitet. PZ-247 Exponate Nah-Infrarot (NIR) Emission mit Lambda-&gt; 700 nm in die erforderliche Wellenl&auml;ngenbereich f&uuml;r maximale Gewebe eindringen. MDA-MB-231-Brust-Tumor-Zellen mit 247 behandelt, der Agent zeigt starke intrazellul&auml;re Fluoreszenz mit einer Emission maximale, 704 nm, gibt an, dass es innerhalb einer hydrophoben Mikromilieu lokalisiert. PZ-247 zeigt assoziieren mit dem lipophilen Kern des LDL und zellul&auml;re Eintrag in erster Linie durch Rezeptor-vermittelte Endozytose ansammeln in Lysosomen zu unterziehen. Erste in-vivo-Studien zeigen, dass 247 pr&auml;ferenziellen Anh&auml;ufung und Speicherung in den Zellen von MDA-MB-231-Tumoren bei M&auml;usen, wodurch NIR optische imaging mit hervorragendem Kontrast zwischen Tumor und umgebende Gewebe unter die Haut implantiert Exponate. Die Intensit&auml;t der Fluoreszenz von 247 innerhalb der Tumor-Erh&ouml;hungen &uuml;ber Zeit bis zu 48 Stunden nach der Injektion vermutlich durch die Beschlagnahme der zirkulierenden 247/LDL komplexe durch das Tumorgewebe. Die Notwendigkeit f&uuml;r Cholesterin und LDL, in hohem Grade proliferative Tumorzellen &uuml;ber nontumorigenic Zellen erh&ouml;ht ist, hat 247 m&ouml;gliche Anwendung f&uuml;r solche Tumoren.

Chirale Porphyrazine nahe-IR optische Bildgebung Agent ausstellen pr&auml;ferenziellen Tumor Akkumulation.

&#x624B;&#x6027; porphyrazine (pz) H(2)[pz(trans-A(2)B(2))] (247)&#xFF0C;&#x7F16;&#x5199;&#x4E86;&#x8BE5;&#x5C55;&#x54C1;&#x4F18;&#x60E0;&#x4F53;&#x5185;&#x79EF;&#x7D2F;&#x5728;&#x7EC6;&#x80DE;&#x4E2D;&#x7684;&#x80BF;&#x7624;&#x3002;Pz 247 &#x5C55;&#x54C1;&#x4E0E; lambda &gt; 700 nm &#x7684;&#x6700;&#x5927;&#x7EC4;&#x7EC7;&#x6E17;&#x900F;&#x6240;&#x9700;&#x7684;&#x6CE2;&#x957F;&#x8303;&#x56F4;&#x5185;&#x7684;&#x8FD1;&#x7EA2;&#x5916; (NIR) &#x6392;&#x653E;&#x3002;&#x5F53;&#x4E19;&#x4E8C;&#x919B;-MB-231 &#x4E73;&#x817A;&#x80BF;&#x7624;&#x7EC6;&#x80DE;&#x7684;&#x6CBB;&#x7597; 247 &#x65F6;&#xFF0C;&#x4EE3;&#x7406;&#x5C06;&#x663E;&#x793A;&#x4E0E;&#x6392;&#x653E;&#x91CF;&#x6700;&#x5927;&#xFF0C;704 &#x5F3A;&#x80DE;&#x5185;&#x7684;&#x8367;&#x5149; nm&#xFF0C;&#x8FD9;&#x8868;&#x660E;&#x5B83;&#x672C;&#x5730;&#x5316;&#x758F;&#x6C34;&#x7684;&#x5FAE;&#x73AF;&#x5883;&#x5185;&#x3002;Pz 247 &#x663E;&#x793A;&#x5173;&#x8054;&#x7684;&#x4F4E;&#x5BC6;&#x5EA6;&#x8102;&#x86CB;&#x767D;&#x7684;&#x4EB2;&#x8102;&#x6027;&#x6838;&#x5FC3;&#x5E76;&#x8FDB;&#x884C;&#x7EC6;&#x80DE;&#x8FDB;&#x5165;&#x4E3B;&#x8981;&#x662F;&#x901A;&#x8FC7;&#x53D7;&#x4F53;&#x4ECB;&#x5BFC;&#x7684;&#x5185;&#x541E;&#x4F5C;&#x7528;&#x5728;&#x6EB6;&#x9176;&#x4F53;&#x4E2D;&#x79EF;&#x7D2F;&#x3002;&#x4F53;&#x5185;&#x7684;&#x521D;&#x6B65;&#x7814;&#x7A76;&#x663E;&#x793A; 247 &#x5C55;&#x54C1;&#x4F18;&#x60E0;&#x79EF;&#x7D2F;&#x548C;&#x4FDD;&#x7559;&#x5728;&#x76AE;&#x4E0B;&#x690D;&#x5165;&#x8001;&#x9F20;&#x4F53;&#x5185;&#xFF0C;&#x4ECE;&#x800C;&#x4F7F;&#x5149;&#x5B66;&#x6210;&#x50CF;&#x4E0E;&#x51FA;&#x8272;&#x7684;&#x5BF9;&#x6BD4;&#x5EA6;&#xFF0C;&#x80BF;&#x7624;&#x4E0E;&#x5468;&#x56F4;&#x7EC4;&#x7EC7;&#x4E4B;&#x95F4;&#x7684;&#x8FD1;&#x7EA2;&#x5916;&#x5149;&#x8C31;&#x7684;&#x4E19;&#x4E8C;&#x919B;-MB-231 &#x80BF;&#x7624;&#x7684;&#x7EC6;&#x80DE;&#x4E2D;&#x3002;247 &#x5185;&#x80BF;&#x7624;&#x4FF1;&#x589E;&#x7684;&#x8367;&#x5149;&#x5F3A;&#x5EA6;&#x5927;&#x6982;&#x662F;&#x56E0;&#x4E3A;&#x87AF;&#x5408;&#x7684;&#x5FAA;&#x73AF; 247/LDL &#x590D;&#x6742;&#x7684;&#x80BF;&#x7624;&#x7EC4;&#x7EC7;&#x6CE8;&#x5C04;&#x540E;&#x7684;&#x65F6;&#x95F4;&#x6700;&#x591A;&#x4E3A; 48 &#x5C0F;&#x65F6;&#x3002;&#x5982;&#x9700;&#x8981;&#x80C6;&#x56FA;&#x9187;&#xFF0C;&#x548C;&#x56E0;&#x6B64;&#x4F4E;&#x5BC6;&#x5EA6;&#x8102;&#x86CB;&#x767D;&#xFF0C;&#x5728;&#x6B63;&#x5E38;&#x7EC6;&#x80DE;&#x9AD8;&#x5EA6;&#x589E;&#x6B96;&#x7684;&#x80BF;&#x7624;&#x7EC6;&#x80DE;&#x5728;&#x5347;&#x9AD8;&#xFF0C;247 &#x5DF2;&#x4E3A;&#x6240;&#x6709;&#x8FD9;&#x7C7B;&#x80BF;&#x7624;&#x7684;&#x6F5C;&#x5728;&#x5E94;&#x7528;&#x3002;

&#x624B;&#x6027; porphyrazine &#x8FD1;&#x7EA2;&#x5916;&#x5149;&#x5B66;&#x6210;&#x50CF;&#x5242;&#x53C2;&#x5C55;&#x4F18;&#x60E0;&#x80BF;&#x7624;&#x79EF;&#x7D2F;&#x3002;

A chiral porphyrazine (pz), H(2)[pz(trans-A(2)B(2))] (247), has been prepared that exhibits preferential in vivo accumulation in the cells of tumors. Pz 247 exhibits near-infrared (NIR) emission with lambda > 700 nm in the required wavelength range for maximum tissue penetration. When MDA-MB-231 breast tumor cells are treated with 247, the agent shows strong intracellular fluorescence with an emission maximum, 704 nm, which indicates that it localizes within a hydrophobic microenvironment. Pz 247 is shown to associate with the lipophilic core of LDL and undergo cellular entry primarily through receptor-mediated endocytosis accumulating in lysosomes. Preliminary in vivo studies show that 247 exhibits preferential accumulation and retention in the cells of MDA-MB-231 tumors subcutaneously implanted in mice, thereby enabling NIR optical imaging with excellent contrast between tumor and surrounding tissue. The intensity of fluorescence from 247 within the tumor increases over time up to 48 h after injection presumably due to the sequestration of circulating 247/LDL complex by the tumor tissue. As the need for cholesterol, and thus LDL, is elevated in highly proliferative tumor cells over nontumorigenic cells, 247 has potential application for all such tumors.

Chiral porphyrazine near-IR optical imaging agent exhibiting preferential tumor accumulation.

Un porphyrazine chiral (pz), H (2) [pz (trans-A (2) B (2))] (247), a &#xE9;t&#xE9; pr&#xE9;par&#xE9; dans l&#x26;#39;accumulation des expositions pr&#xE9;f&#xE9;rentiels in vivo dans les cellules de tumeurs. Pz 247 expositions dans le proche infrarouge (NIR) des &#xE9;missions avec lambda&#x26;gt; 700 nm dans la gamme de longueur d&#x26;#39;onde n&#xE9;cessaire pour la p&#xE9;n&#xE9;tration tissulaire maximale. Lorsque des cellules tumorales MDA-MB-231 du sein sont trait&#xE9;s avec 247, l&#x26;#39;agent pr&#xE9;sente une forte fluorescence intracellulaire avec un maximum d&#x26;#39;&#xE9;mission, 704 nm, ce qui indique qu&#x26;#39;il localise dans un micro-environnement hydrophobe. Pz 247 est montr&#xE9; &#xE0; associer avec le noyau lipophile de LDL et de subir l&#x26;#39;entr&#xE9;e cellulaire principalement par endocytose s&#x26;#39;accumulant dans les lysosomes. Pr&#xE9;liminaire des &#xE9;tudes in vivo montrent que l&#x26;#39;accumulation de 247 expositions pr&#xE9;f&#xE9;rentiel et la r&#xE9;tention dans les cellules de MDA-MB-231 tumeurs implant&#xE9;es chez des souris par voie sous cutan&#xE9;e, ce qui permet l&#x26;#39;imagerie optique NIR avec un excellent contraste entre la tumeur et des tissus environnants. L&#x26;#39;intensit&#xE9; de fluorescence &#xE0; partir de 247 au sein de la tumeur augmente au fil du temps jusqu&#x26;#39;&#xE0; 48 h apr&#xE8;s l&#x26;#39;injection sans doute en raison de la s&#xE9;questration de circulation complexe 247/LDL par le tissu tumoral. Comme la n&#xE9;cessit&#xE9; pour le cholest&#xE9;rol, LDL et donc, est tr&#xE8;s &#xE9;lev&#xE9;e dans les cellules tumorales prolif&#xE9;rantes sur les cellules non tumorig&#xE8;nes, 247 dispose d&#x26;#39;applications potentielles pour toutes les tumeurs telles.

Chirale porphyrazine Proche-IR agent d&#x26;#39;imagerie optique Exposer accumulation tumorale pr&#xE9;f&#xE9;rentiel

Un porphyrazine chirale (pz), H(2)[pz(trans-A(2)B(2))] (247), &egrave; stato preparato che accumulo in vivo preferenziale di mostre nelle cellule dei tumori. 247 Pz presenta emissione di vicino infrarosso (NIR) con lambda &amp;gt; 700 nm nell&#39;intervallo di lunghezza d&#39;onda richiesta per penetrazione massima tessuto. Quando le cellule tumorali di mammella MDA-MB-231 sono trattate con 247, l&#39;agente Mostra forte fluorescenza intracellulare con un&#39;emissione massima, 704 nm, che indica che localizza all&#39;interno di un microambiente idrofobico. 247 Pz &egrave; dimostrato associato con il nucleo lipofilo di LDL e sottoposti a voce cellulare principalmente tramite endocitosi mediata dal recettore accumulo nei lisosomi. Preliminari studi in vivo dimostrano che 247 esibisce preferenziale accumulo e ritenzione nelle cellule dei tumori MDA-MB-231 per via sottocutanea impiantati nei topi, consentendo in tal modo NIR optical imaging con ottimo contrasto tra il tumore e il tessuto circostante. L&#39;intensit&agrave; di fluorescenza da 247 entro il tumore aumenta oltre il tempo fino a 48 h dopo l&#39;iniezione, presumibilmente a causa del sequestro di 247/LDL complesso in circolazione dal tessuto tumorale. Come il bisogno di colesterolo e cos&igrave; LDL, &egrave; elevato nelle cellule tumorali altamente proliferative sopra cellule nontumorigenic, 247 ha potenziale applicazione per tutti i tali tumori.

Porphyrazine chirale vicino-IR optical imaging agente espositrici accumulo preferenziale del tumore.

&#x30AD;&#x30E9;&#x30EB;&#x30DD;&#x30EB;&#x30D5;&#x30A3;&#x30E9;&#x30B8;&#x30F3;&#xFF08;PZ&#xFF09;&#x3001;H&#xFF08;2&#xFF09;[PZ&#xFF08;&#x30C8;&#x30E9;&#x30F3;&#x30B9;-A&#xFF08;2&#xFF09;B&#xFF08;2&#xFF09;&#xFF09;]&#xFF08;247&#xFF09;&#x3001;&#x4F5C;&#x6210;&#x3055;&#x308C;&#x305F;&#x305D;&#x306E;&#x816B;&#x760D;&#x306E;&#x7D30;&#x80DE;&#x304C;in vivo&#x3067;&#x306E;&#x84C4;&#x7A4D;&#x306E;&#x512A;&#x9047;&#x5C55;&#x793A;&#x3002;&#x30E9;&#x30E0;&#x30C0;&#x3068;&#x8FD1;&#x8D64;&#x5916;&#xFF08;NIR&#xFF09;&#x6392;&#x51FA;PZ 247&#x5C55;&#x793A;&#x6700;&#x5927;&#x306E;&#x7D44;&#x7E54;&#x3078;&#x306E;&#x6D78;&#x900F;&#x306E;&#x305F;&#x3081;&#x306B;&#x5FC5;&#x8981;&#x306A;&#x6CE2;&#x9577;&#x7BC4;&#x56F2;&#x306E;&#x26;gt; 700 nm&#x3067;&#x3042;&#x308B;&#x3002; MDA-MB-231&#x4E73;&#x764C;&#x7D30;&#x80DE;&#x306F;247&#x3067;&#x51E6;&#x7406;&#x3055;&#x308C;&#x3066;&#x3044;&#x308B;&#x5834;&#x5408;&#x3001;&#x30A8;&#x30FC;&#x30B8;&#x30A7;&#x30F3;&#x30C8;&#x306F;&#x3001;&#x758E;&#x6C34;&#x6027;&#x306E;&#x5FAE;&#x5C0F;&#x74B0;&#x5883;&#x5185;&#x306B;&#x5C40;&#x5728;&#x3059;&#x308B;&#x3053;&#x3068;&#x3092;&#x793A;&#x3057;&#x3066;&#x767A;&#x5149;&#x6975;&#x5927;&#x3001;704 nm&#x3067;&#x3042;&#x308A;&#x3001;&#x3068;&#x306E;&#x5F37;&#x529B;&#x306A;&#x7D30;&#x80DE;&#x5185;&#x86CD;&#x5149;&#x3092;&#x793A;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002; PZ 247 LDL&#x306E;&#x89AA;&#x6CB9;&#x6027;&#x30B3;&#x30A2;&#x306B;&#x95A2;&#x9023;&#x4ED8;&#x3051;&#x3089;&#x308C;&#x3001;&#x4E3B;&#x306B;&#x30EA;&#x30BD;&#x30BD;&#x30FC;&#x30E0;&#x306B;&#x84C4;&#x7A4D;&#x3059;&#x308B;&#x53D7;&#x5BB9;&#x4F53;&#x3092;&#x4ECB;&#x3057;&#x305F;&#x30A8;&#x30F3;&#x30C9;&#x30B5;&#x30A4;&#x30C8;&#x30FC;&#x30B7;&#x30B9;&#x3092;&#x4ECB;&#x3057;&#x3066;&#x7D30;&#x80DE;&#x306E;&#x30A8;&#x30F3;&#x30C8;&#x30EA;&#x3092;&#x53D7;&#x3051;&#x308B;&#x3053;&#x3068;&#x304C;&#x793A;&#x3055;&#x308C;&#x3066;&#x3044;&#x308B;&#x3002; in vivo&#x8A66;&#x9A13;&#x306E;&#x4E88;&#x5099;&#x306F;&#x3001;&#x76AE;&#x4E0B;&#x306B;&#x30DE;&#x30A6;&#x30B9;&#x306B;&#x79FB;&#x690D;MDA-MB-231&#x816B;&#x760D;&#x306E;&#x7D30;&#x80DE;&#x306B;&#x3042;&#x308B;247&#x8ED2;&#x306E;&#x5C55;&#x793A;&#x512A;&#x9047;&#x84C4;&#x7A4D;&#x3068;&#x4FDD;&#x6301;&#x3057;&#x3001;&#x305D;&#x308C;&#x306B;&#x3088;&#x3063;&#x3066;&#x816B;&#x760D;&#x3068;&#x5468;&#x56F2;&#x7D44;&#x7E54;&#x3068;&#x306E;&#x9593;&#x306E;&#x512A;&#x308C;&#x305F;&#x30B3;&#x30F3;&#x30C8;&#x30E9;&#x30B9;&#x30C8;&#x3092;&#x6301;&#x3064;&#x8FD1;&#x8D64;&#x5916;&#x5149;&#x30A4;&#x30E1;&#x30FC;&#x30B8;&#x30F3;&#x30B0;&#x3092;&#x53EF;&#x80FD;&#x306B;&#x3059;&#x308B;&#x3053;&#x3068;&#x3092;&#x793A;&#x3057;&#x3066;&#x3044;&#x308B;&#x3002;&#x816B;&#x760D;&#x7D44;&#x7E54;&#x3067;247/LDL&#x8907;&#x96D1;&#x306A;&#x5FAA;&#x74B0;&#x306E;&#x9694;&#x96E2;&#x306B;&#x3088;&#x308B;&#x3082;&#x306E;&#x3067;&#x3042;&#x308D;&#x3046;&#x6CE8;&#x5C04;&#x5F8C;&#x6700;&#x5927;48&#x6642;&#x9593;&#x307E;&#x3067;&#x7D4C;&#x6642;&#x7684;&#x816B;&#x760D;&#x5897;&#x5927;&#x3059;&#x308B;&#x4E2D;&#x3067;247&#x304B;&#x3089;&#x306E;&#x86CD;&#x5149;&#x5F37;&#x5EA6;&#x3002; LDL&#x30B3;&#x30EC;&#x30B9;&#x30C6;&#x30ED;&#x30FC;&#x30EB;&#x306E;&#x5FC5;&#x8981;&#x6027;&#x3068;&#x3057;&#x3066;&#x3001;&#x3057;&#x305F;&#x304C;&#x3063;&#x3066;&#x3001;247&#x306F;&#x3059;&#x3079;&#x3066;&#x306E;&#x305D;&#x306E;&#x3088;&#x3046;&#x306A;&#x816B;&#x760D;&#x306B;&#x5BFE;&#x3059;&#x308B;&#x6F5C;&#x5728;&#x7684;&#x306A;&#x30A2;&#x30D7;&#x30EA;&#x30B1;&#x30FC;&#x30B7;&#x30E7;&#x30F3;&#x304C;&#x3042;&#x308A;&#x307E;&#x3059;&#x304C;&#x3001;&#x975E;&#x816B;&#x760D;&#x5F62;&#x6210;&#x6027;&#x7D30;&#x80DE;&#x3092;&#x4ECB;&#x3057;&#x3066;&#x9AD8;&#x5EA6;&#x306B;&#x5897;&#x6B96;&#x3059;&#x308B;&#x816B;&#x760D;&#x7D30;&#x80DE;&#x3067;&#x4E0A;&#x6607;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;

&#x512A;&#x9047;&#x816B;&#x760D;&#x306E;&#x84C4;&#x7A4D;&#x3092;&#x5C55;&#x793A;&#x30AD;&#x30E9;&#x30EB;&#x30DD;&#x30EB;&#x30D5;&#x30A3;&#x30E9;&#x30B8;&#x30F3;&#x8FD1;&#x8D64;&#x5916;&#x5149;&#x30A4;&#x30E1;&#x30FC;&#x30B8;&#x30F3;&#x30B0;&#x30A8;&#x30FC;&#x30B8;&#x30A7;&#x30F3;&#x30C8;

&#xD0A4; &#xB784; porphyrazine (&#xD3EC; &#xD150; &#xCC28;), H(2)[pz(trans-A(2)B(2))] (247), &#xADF8; &#xC804;&#xC2DC; &#xC6B0;&#xC120; &#xC0DD;&#xCCB4; &#xC870;&#xAC74; &#xC140;&#xC5D0;&#xC11C;&#xC758; &#xCD95;&#xC801; &#xC885;&#xC591; &#xC900;&#xBE44; &#xB418;&#xC5C8;&#xC2B5;&#xB2C8;&#xB2E4;. Pz 247 &#xC804;&#xC2DC; &#xCD5C;&#xB300; &#xC870;&#xC9C1; &#xCE68;&#xD22C;&#xB97C; &#xC704;&#xD55C; &#xD544;&#xC694;&#xD55C; &#xD30C;&#xC7A5; &#xBC94;&#xC704;&#xC5D0;&#xC11C; &#xB78C;&#xB2E4; &amp;gt; 700 nm&#xC640; &#xADFC; &#xC801;&#xC678;&#xC120; (NIR) &#xBC29;&#xCD9C;. MDA-MB-231 &#xC720; &#xBC29; &#xC885;&#xC591; &#xC138;&#xD3EC;&#xB294; 247&#xC640; &#xD568;&#xAED8; &#xCC98;&#xB9AC; &#xB429;&#xB2C8;&#xB2E4;, &#xC5D0;&#xC774;&#xC804;&#xD2B8;&#xB294; &#xBC29;&#xCD9C; &#xCD5C;&#xB300;, 704&#xC640; &#xAC15;&#xD55C; &#xC138;&#xD3EC;&#xB0B4; &#xD615;&#xAD11; &#xD45C;&#xC2DC; nm, &#xC18C;&#xC218; &#xC131; microenvironment &#xB0B4;&#xC5D0;&#xC11C; localizes &#xB098;&#xD0C0;&#xB0C5;&#xB2C8;&#xB2E4;. Pz 247 LDL&#xC758; &#xBE14;&#xB808;&#xBBF8;&#xC26C; &#xCF54;&#xC5B4;&#xC640; &#xC5F0;&#xACB0; &#xD558; &#xC5EC; &#xC8FC;&#xB85C; &#xC218;&#xC6A9; &#xCCB4;-&#xC911;&#xC7AC; endocytosis &#xB9AC;&#xC18C;&#xC880; &#xCD95;&#xC801;&#xC744; &#xD1B5;&#xD574; &#xC140;&#xB8F0;&#xB7EC; &#xD56D;&#xBAA9;&#xC744; &#xBC1B; &#xB2E4; &#xD45C;&#xC2DC; &#xB429;&#xB2C8;&#xB2E4;. &#xC608;&#xBE44; in vivo &#xC5F0;&#xAD6C; 247 &#xD2B9;&#xD61C; &#xCD95;&#xC801;&#xACFC; subcutaneously &#xBBC0;&#xB85C; NIR &#xC885;&#xC591;&#xACFC; &#xC8FC;&#xC704; &#xC870;&#xC9C1; &#xAC04;&#xC758; &#xB6F0;&#xC5B4;&#xB09C; &#xBA85;&#xC554;&#xBE44;&#xC640; &#xC774;&#xBBF8;&#xC9D5; &#xAD11;&#xD559; &#xC950;&#xC5D0; &#xC774;&#xC2DD; &#xD558;&#xB294; MDA-MB-231 &#xC885;&#xC591; &#xC138;&#xD3EC;&#xC5D0; &#xBCF4;&#xC874; &#xC804;&#xC2DC; &#xBCF4;&#xC5EC;&#xC90D;&#xB2C8;&#xB2E4;. &#xD1B5;&#xD574; &#xC885;&#xC591; &#xC99D;&#xAC00; &#xB0B4; 247&#xC5D0;&#xC11C; &#xD615;&#xAD11;&#xC758; &#xAC15;&#xB3C4; &#xC885;&#xC591; &#xC870;&#xC9C1;&#xC5D0; &#xC758;&#xD574; 247/LDL &#xBCF5;&#xC7A1; &#xD55C; &#xC21C;&#xD658;&#xC758; &#xACA9;&#xB9AC; &#xB54C;&#xBB38;&#xC5D0; &#xC544;&#xB9C8;&#xB3C4; &#xC8FC;&#xC0AC; &#xD6C4; 48 h &#xC2DC;&#xAC04;. &#xCF5C;&#xB808;&#xC2A4;&#xD14C;&#xB864; &#xBC0F; LDL, &#xB530;&#xB77C;&#xC11C;&#xC5D0; &#xB300; &#xD55C; &#xD544;&#xC694;&#xC131;&#xC744; &#xB192;&#xC740; &#xC99D;&#xC2DD; &#xC885;&#xC591; &#xC138;&#xD3EC;&#xC5D0;&#xC11C; nontumorigenic &#xC140;&#xC744; &#xD1B5;&#xD574; &#xC0C1;&#xC2B9;&#xC73C;&#xB85C; 247 &#xAC19;&#xC740; &#xC885;&#xC591;&#xC5D0; &#xB300; &#xD55C; &#xC7A0;&#xC7AC;&#xC801;&#xC778; &#xC751;&#xC6A9; &#xD504;&#xB85C;&#xADF8;&#xB7A8;&#xC740;.

&#xD0A4; &#xB784; porphyrazine &#xADFC;&#xCC98;-&#xC801;&#xC678;&#xC120; &#xAD11;&#xD559; &#xC774;&#xBBF8;&#xC9D5; &#xC5D0;&#xC774;&#xC804;&#xD2B8; &#xC6B0;&#xC120; &#xC885;&#xC591; &#xCD95;&#xC801;&#xC744; &#xC804;&#xC2DC;.

Un porfirazina quiral (PZ), H (2) [PZ (trans-A (2) B (2))] (247), que se ha preparado exposiciones preferentes en la acumulaci&#xF3;n in vivo en las c&#xE9;lulas de tumores. Pz 247 exposiciones en el infrarrojo cercano (NIR) de las emisiones con lambda&#x26;gt; 700 nm en el rango de longitud de onda necesaria para la m&#xE1;xima penetraci&#xF3;n en el tejido. Cuando MDA-MB-231 de mama las c&#xE9;lulas tumorales se tratan con 247, el agente muestra fuerte fluorescencia intracelular con un m&#xE1;ximo de emisi&#xF3;n, 704 nm, lo que indica que se localiza dentro de un microambiente hidr&#xF3;fobo. Pz 247 se muestra a asociar con el n&#xFA;cleo lip&#xF3;filo de las LDL y se someten a la entrada celular, principalmente a trav&#xE9;s de endocitosis mediada por receptor acumulaci&#xF3;n en los lisosomas. Los estudios preliminares in vivo muestran que la acumulaci&#xF3;n de 247 exposiciones preferencial y la retenci&#xF3;n en las c&#xE9;lulas de MDA-MB-231 tumores implantados en ratones por v&#xED;a subcut&#xE1;nea, permitiendo as&#xED; la formaci&#xF3;n de im&#xE1;genes NIR &#xF3;ptica con excelente contraste entre el tumor y el tejido circundante. La intensidad de fluorescencia de 247 dentro de los tumor aumenta con el tiempo hasta 48 h despu&#xE9;s de la inyecci&#xF3;n, presumiblemente debido a la retenci&#xF3;n del complejo circulante 247/LDL por el tejido tumoral. Como la necesidad de colesterol, LDL y por lo tanto, se eleva en las c&#xE9;lulas tumorales altamente proliferativos sobre las c&#xE9;lulas nontumorigenic, 247 tiene una aplicaci&#xF3;n potencial para todo tipo de tumores.

Porfirazina quiral infrarrojo cercano agente de imagen &#xF3;ptico Exponer acumulaci&#xF3;n preferencial del tumor

Molecular imaging provides spatial and temporal information on cellular changes that occur during development and in disease. MRI and optical imaging of reporter genes allows for the visualization of promoter activity, protein-protein interactions, protein stability and the tracking of individual proteins and cells. Reporter genes can be genetically encoded in transgenic animals or detected through the administration of an exogenous contrast agent. Advances in molecular imaging of reporter genes have led to the development of imaging probes that detect changes in endogenous cellular changes. The ability to use contrast agents coupled with functional information on cellular events will allow for sensitive assessment of individual patient therapies, leading to an accurately tailored treatment regimen.

Molecular imaging of in vivo gene expression.

We describe the use of Co(III) Schiff base-DNA conjugates, a versatile class of research tools that target C2H2 transcription factors, to inhibit the Hedgehog (Hh) pathway. In developing mammalian embryos, Hh signaling is critical for the formation and development of many tissues and organs. Inappropriate activation of the Hedgehog (Hh) pathway has been implicated in a variety of cancers including medulloblastomas and basal cell carcinomas. It is well-known that Hh regulates the activity of the Gli family of C2H2 zinc finger transcription factors in mammals. In Drosophila the function of the Gli proteins is performed by a single transcription factor with an identical DNA binding consensus sequence, Cubitus Interruptus (Ci). We have demonstrated previously that conjugation of a specific 17 base-pair oligonucleotide to a Co(III) Schiff base complex results in a targeted inhibitor of the Snail family C2H2 zinc finger transcription factors. Modification of the oligonucleotide sequence in the Co(III) Schiff base-DNA conjugate to that of Ci's consensus sequence (Co(III)-Ci) generates an equally selective inhibitor of Ci. Co(III)-Ci irreversibly binds the Ci zinc finger domain and prevents it from binding DNA in vitro. In a Ci responsive tissue culture reporter gene assay, Co(III)-Ci reduces the transcriptional activity of Ci in a concentration dependent manner. In addition, injection of wild-type Drosophila embryos with Co(III)-Ci phenocopies a Ci loss of function phenotype, demonstrating effectiveness in vivo. This study provides evidence that Co(III) Schiff base-DNA conjugates are a versatile class of specific and potent tools for studying zinc finger domain proteins and have potential applications as customizable anticancer therapeutics.

Specific inhibition of the transcription factor Ci by a cobalt(III) Schiff base-DNA conjugate.

Les auteurs d&eacute;crivent l&#39;utilisation de conjugu&eacute;s d&#39;ADN-base de Schiff Co (III), une classe polyvalente d&#39;outils de recherche qui ciblent les facteurs de transcription C2H2, inhibe la voie Hedgehog (Hh). Dans le d&eacute;veloppement des embryons chez les mammif&egrave;res, la signalisation de Hh est critique pour la formation et le d&eacute;veloppement de nombreux tissus et organes. Activation inappropri&eacute;e de la voie de h&eacute;risson (Hh) a &eacute;t&eacute; impliqu&eacute;e dans une vari&eacute;t&eacute; de cancers dont des m&eacute;dulloblastomes et des carcinomes baso-cellulaires. Il est bien connu que les Hh r&eacute;glemente l&#39;activit&eacute; de la famille de Gli de facteurs de transcription doigt C2H2 zinc chez les mammif&egrave;res. Chez la drosophile, la fonction des prot&eacute;ines Gli est exerc&eacute;e par un facteur de transcription unique avec une s&eacute;quence de consensus de liaison ADN identique, Cubitus Interruptus (Ci). Nous avons d&eacute;montr&eacute; pr&eacute;c&eacute;demment que la conjugaison d&#39;un oligonucl&eacute;otide sp&eacute;cifique de paires de bases 17 &agrave; un co (III) Schiff base complexe g&eacute;n&egrave;re un inhibiteur cibl&eacute; des facteurs de transcription du doigt de zinc C2H2 familles escargot. Modification de la s&eacute;quence dans le conjugu&eacute; d&#39;ADN-base de Schiff Co (III) &agrave; celle de la s&eacute;quence consensus de Ci (Co(III)-Ci) g&eacute;n&egrave;re un inhibiteur s&eacute;lectif &eacute;galement de Ci. Co (III)-Ci a irr&eacute;versiblement lie du domaine de doigt de zinc de Ci et l&#39;emp&ecirc;che de lier l&#39;ADN in vitro. Dans un essai de g&egrave;ne reporter des vitroplants sensible-Ci, Co (III)-Ci a r&eacute;duit l&#39;activit&eacute; transcriptionnelle de Ci de mani&egrave;re d&eacute;pendante de la concentration. En outre, injection d&#39;embryons de drosophile sauvage avec Co (III)-Ci ph&eacute;nocopies un Ci perte du ph&eacute;notype de fonction, ce qui d&eacute;montre l&#39;efficacit&eacute; in vivo. Cette &eacute;tude d&eacute;montre que les conjugu&eacute;s d&#39;ADN-base de Schiff Co (III) sont une classe polyvalente d&#39;outils pr&eacute;cis et puissants pour l&#39;&eacute;tude des prot&eacute;ines domaine en doigt de zinc et ont des applications potentielles comme th&eacute;rapeutique anticanc&eacute;reuse personnalisable.

Inhibition sp&eacute;cifique du facteur de transcription Ci par un conjugu&eacute; de cobalt (III) ADN-base de Schiff.

Descriviamo l&#39;uso dei coniugati di base-DNA Co(III) Schiff, una classe versatile di strumenti di ricerca che l&#39;obiettivo di fattori di trascrizione C2H2, per inibire il pathway Hedgehog (Hh). Lo sviluppo di embrioni dei mammiferi, Hh di segnalazione &egrave; fondamentale per la formazione e lo sviluppo di molti tessuti e organi. Attivazione inappropriata del percorso Hedgehog (Hh) &egrave; stato implicato in una variet&agrave; di tumori tra cui medulloblastomi e carcinomi a cellule basali. &Egrave; noto che Hh regola l&#39;attivit&agrave; della famiglia Gli C2H2 zinco dito fattori di trascrizione nei mammiferi. In Drosophila viene eseguita la funzione delle proteine Gli da un fattore di trascrizione singolo con un&#39;identica sequenza consenso del DNA binding, Cubitus Interruptus (Ci). Abbiamo dimostrato in precedenza che coniugazione di un oligonucleotide di base-coppia 17 specifici per un Co(III) Schiff base complessa provoca un inibitore mirato il lumaca famiglia C2H2 zinco dito fattori di trascrizione. Modifica della sequenza di oligonucleotidi nel coniugato di base-DNA Co(III) Schiff a quella della sequenza di consenso di Ci (Co(III)-Ci) genera un altrettanto selettivo inibitore di Ci. Co (III)-Ci irreversibilmente si lega al dominio del dito di zinco Ci e impedisce di legare il DNA in vitro. In un saggio di gene di reporter del coltura del tessuto reattivo Ci, Co (III)-Ci riduce l&#39;attivit&agrave; trascrizionale di Ci in modo dipendente dalla concentrazione. In aggiunta, iniezione di embrioni della drosofila selvaggio-tipo con Co (III)-Ci fenocopie un Ci perdita del fenotipo di funzione, dimostrando l&#39;efficacia in vivo. Questo studio fornisce la prova che coniugati di base-DNA Co(III) Schiff sono una classe versatile di specifici e potenti strumenti per lo studio delle proteine di dominio zinc finger ed hanno applicazioni potenziali come terapie antitumorali personalizzabile.

Inibizione specifica del fattore di trascrizione Ci da un base di Schiff-DNA cobalt(III) coniugato.

Snail family proteins are core EMT (epithelial-mesenchymal transition) regulatory factors that play essential roles in both development and disease processes and have been associated with metastasis in carcinomas. Snail factors are required for the formation of neural crest stem cells in most vertebrate embryos, as well as for the migratory invasive behavior of these cells. Snail factors have recently been linked to the formation of cancer stem cells, and expression of Snail proteins may be associated with tumor recurrence and resistance to chemotherapy and radiotherapy. We report that Co(III)-Ebox is a potent inhibitor of Snail-mediated transcriptional repression in breast cancer cells and in the neural crest of Xenopus. We further show that the activity of Co(III)-Ebox can be modulated by temperature, increasing the utility of this conjugate as a Snail inhibitor in model organisms. We exploit this feature to further delineate the requirements for Snail function during neural crest development, showing that in addition to the roles that Snail factors play in neural crest precursor formation and neural crest EMT/migration, inhibition of Snail function after the onset of neural crest migration leads to a loss of neural crest derived melanocytes. Co(III)-Ebox-mediated inhibition therefore provides a powerful tool for analysing the function of these core EMT factors with unparalleled temporal resolution. Moreover, the potency of Co(III)-Ebox as a Snail inhibitor in breast cancer cells suggests its potential as a therapeutic inhibitor of tumor progression and metastasis.

Targeted inactivation of Snail family EMT regulatory factors by a Co(III)-Ebox conjugate.

Cobalt(III) Schiff base complexes have been used as potent inhibitors of protein function through the coordination to histidine residues essential for activity. The kinetics and thermodynamics of the binding mechanism of Co(acacen)(NH(3))(2)Cl [Co(acacen); where H(2)acacen is bis(acetylacetone)ethylenediimine] enzyme inhibition has been examined through the inactivation of matrix metalloproteinase 2 (MMP-2) protease activity. Co(acacen) is an irreversible inhibitor that exhibits time- and concentration-dependent inactivation of MMP-2. Co(acacen) inhibition of MMP-2 is temperature-dependent, with the inactivation increasing with temperature. Examination of the formation of the transition state for the MMP-2/Co(acacen) complex was determined to have a positive entropy component indicative of greater disorder in the MMP-2/Co(acacen) complex than in the reactants. With further insight into the mechanism of Co(acacen) complexes, Co(III) Schiff base complex protein inactivators can be designed to include features regulating activity and protein specificity. This approach is widely applicable to protein targets that have been identified to have clinical significance, including matrix metalloproteinases. The mechanistic information elucidated here further emphasizes the versatility and utility of Co(III) Schiff base complexes as customizable protein inhibitors.

Kinetics and thermodynamics of irreversible inhibition of matrix metalloproteinase 2 by a Co(III) Schiff base complex.

The kinetic and thermodynamic ligand exchange dynamics are important considerations in the rational design of metal-based therapeutics and therefore, require detailed investigation. Co(III) Schiff base complex derivatives of bis(acetylacetone)ethylenediimine [acacen] have been found to be potent enzyme and transcription factor inhibitors. These complexes undergo solution exchange of labile axial ligands. Upon dissociation, Co(III) irreversibly interacts with specific histidine residues of a protein, and consequently alters structure and causes inhibition. To guide the rational design of next generation agents, understanding the mechanism and dynamics of the ligand exchange process is essential. To investigate the lability, pH stability, and axial ligand exchange of these complexes in the absence of proteins, the pD- and temperature-dependent axial ligand substitution dynamics of a series of N-heterocyclic [Co(acacen)(X)(2)](+) complexes [where X = 2-methylimidazole (2MeIm), 4-methylimidazole (4MeIm), ammine (NH(3)), N-methylimidazole (NMeIm), and pyridine (Py)] were characterized by NMR spectroscopy. The pD stability was shown to be closely related to the nature of the axial ligand with the following trend toward aquation: 2MeIm > NH(3) ≫ 4MeIm > Py > Im > NMeIm. Reaction of each [Co(III)(acacen)(X)(2)](+) derivative with 4MeIm showed formation of a mixed ligand Co(III) intermediate via a dissociative ligand exchange mechanism. The stability of the mixed ligand adduct was directly correlated to the pD-dependent stability of the starting Co(III) Schiff base with respect to [Co(acacen)(4MeIm)(2)](+). Crystal structure analysis of the [Co(acacen)(X)(2)](+) derivatives confirmed the trends in stability observed by NMR spectroscopy. Bond distances between the Co(III) and the axial nitrogen atoms were longest in the 2MeIm derivative as a result of distortion in the planar tetradentate ligand, and this was directly correlated to axial ligand lability and propensity toward exchange.

Axial ligand exchange of N-heterocyclic cobalt(III) Schiff base complexes: molecular structure and NMR solution dynamics.

A bacteria-targeted MR contrast agent, Zn-1, consisting of two Zn-dipicolylamine (Zn-dpa) groups conjugated to a Gd(III) chelate has been synthesized and characterized. In vitro studies with S. aureus and E. coli show that Zn-1 exhibits a significant improvement in bacteria labeling efficiency vs. control. Studies with a structural analogue, Zn-2, indicate that removal of one Zn-dpa moiety dramatically reduces the agent's affinity for bacteria. The ability of Zn-1 to significantly reduce the T 1 of labeled vs. unlabeled bacteria, resulting in enhanced MR image contrast, demonstrates its potential for visualizing bacterial infections in vivo.

Synthesis, Characterization, and in vitro Testing of a Bacteria-Targeted MR Contrast Agent.

Dissemination of tumor cells is an essential step in metastasis. Direct contact between a macrophage, mammalian-enabled (MENA)-overexpressing tumor cell, and endothelial cell [Tumor MicroEnvironment of Metastasis (TMEM)] correlates with metastasis in breast cancer patients. Here we show, using intravital high-resolution two-photon microscopy, that transient vascular permeability and tumor cell intravasation occur simultaneously and exclusively at TMEM. The hyperpermeable nature of tumor vasculature is described as spatially and temporally heterogeneous. Using real-time imaging, we observed that vascular permeability is transient, restricted to the TMEM, and required for tumor cell dissemination. VEGFA signaling from TIE2(hi) TMEM macrophages causes local loss of vascular junctions, transient vascular permeability, and tumor cell intravasation, demonstrating a role for the TMEM within the primary mammary tumor. These data provide insight into the mechanism of tumor cell intravasation and vascular permeability in breast cancer, explaining the value of TMEM density as a predictor of distant metastatic recurrence in patients.

Real-Time Imaging Reveals Local, Transient Vascular Permeability, and Tumor Cell Intravasation Stimulated by TIE2hi Macrophage-Derived VEGFA.

Cobalt(III) Schiff base complexes ([Co(acacen)(L)2](+), where L = NH3) inhibit histidine-containing proteins through dissociative exchange of the labile axial ligands (L). This work investigates axial ligand exchange dynamics of [Co(acacen)(L)2](+) complexes toward the development of protein inhibitors that are activated by external triggers such as light irradiation. We sought to investigate ligand exchange dynamics to design a Co(III) complex that is substitutionally inert under normal physiological conditions for selective activation. Fluorescent imidazoles (C3Im) were prepared as axial ligands in [Co(acacen)(L)2](+) to produce complexes (CoC3Im) that could report on ligand exchange and, thus, complex stability. These fluorescent imidazole reporters guided the design of a new dinuclear Co(III) Schiff base complex containing bridging diimidazole ligands, which exhibits enhanced stability to ligand exchange with competing imidazoles and to hydrolysis within a biologically relevant pH range. These studies inform the design of biocompatible Co(III) Schiff base complexes that can be selectively activated for protein inhibition with spatial and temporal specificity.

Tuning cobalt(III) Schiff base complexes as activated protein inhibitors.

As molecular labels for cells and tissues, fluorescent probes have shaped our understanding of biological structures and processes. However, their capacity for quantitative analysis is limited because photon emission rates from multicolour fluorophores are dissimilar, unstable and often unpredictable, which obscures correlations between measured fluorescence and molecular concentration. Here we introduce a new class of light-emitting quantum dots with tunable and equalized fluorescence brightness across a broad range of colours. The key feature is independent tunability of emission wavelength, extinction coefficient and quantum yield through distinct structural domains in the nanocrystal. Precise tuning eliminates a 100-fold red-to-green brightness mismatch of size-tuned quantum dots at the ensemble and single-particle levels, which substantially improves quantitative imaging accuracy in biological tissue. We anticipate that these materials engineering principles will vastly expand the optical engineering landscape of fluorescent probes, facilitate quantitative multicolour imaging in living tissue and improve colour tuning in light-emitting devices.

Allison S. Harney

Department of Anatomy & Structural Biology,
Department of Radiology,
Gruss-Lipper Biophotonics Center,
Integrated Imaging Program

Albert Einstein College of Medicine

Extended Time-lapse Intravital Imaging of Real-time Multicellular Dynamics in the Tumor Microenvironment

Allison S. Harney

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Anatomy & Structural Biology,Department of Radiology,Gruss-Lipper Biophotonics Center,Integrated Imaging Program

Albert Einstein College of Medicine

Extended Time-lapse Intravital Imaging of Real-time Multicellular Dynamics in the Tumor Microenvironment

Department of Anatomy & Structural Biology,
Department of Radiology,
Gruss-Lipper Biophotonics Center,
Integrated Imaging Program