Name: a syngeneic mouse model of metastatic renal cell carcinoma for quantitative and longitudinal assessment of preclinical therapies
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Description: Watch this Scientific Journal Video about A Syngeneic Mouse Model of Metastatic Renal Cell Carcinoma for Quantitative and Longitudinal Assessment of Preclinical Therapies at JoVE.com

Dette arbejde blev støttet af tilskud fra National Cancer Institute (R15CA173657 til AW og R01CA109446 til GTS), Simmons Cancer Institute (til AW), og University of Minnestoa forcere 4 Nyre Cancer Foundation (til GTS). Vi takker Dr. Kristin Anderson for at få hjælp med immunofluorescens.

Følgende protokol beskriver en eksperimentel procedure til induktion og overvågning af vækst af eksperimentelle ortotope renale tumorer og eventuelle spontane metastaser. Alle de følgende procedurer er udført i overensstemmelse med de institutionelle politikker og godkendte procedurer for human brug af forsøgsdyr. 1. Vedligeholdelse af cellelinier <ol><li> Opretholde murine renal adenocarcinom cellelinie, Renca, i Roswell Park Memorial Institute (RPMI) 1640 medium suppleret med 10% føtalt kalveserum (FC...

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Vi præsenterer en protokol for en ortotopisk RCC musemodel. Implantation af muse renale adenocarcinom tumorceller i musen nyre tilvejebringer en klinisk relevant model for mRCC. Denne model resulterer i en primær tumor i nyren og fjernmetastaser i lungerne, som begge er kendetegnende for fremskreden RCC. Beskrivelsen er præsenteret heri er specifik for Renca cellelinien i immunkompetente Balb / c-mus. Under anvendelse af denne orthotopisk model har vi vist, at Renca tumorer reagerer på immunterapi, hvilket viser anv...

Nyrecellecancer (RCC) tegner sig for hovedparten af maligne nyre neoplasmer og omkring 3% af alle voksne maligniteter verdensplan 1, 2. På grund af manglende symptomer, fejldiagnoser og utilstrækkelige screening værktøjer til RCC, vil næsten 30% af patienterne præsentere med metastatisk RCC (mRCC) på tidspunktet for diagnosen, med yderligere 20-30% af patienterne udvikler sig til et metastatisk stadie 2. Disse sager resulterer i ~ 13.500 dødsfald årligt 1, 3. Selvom tidligere påvisning og behandling af primær RCC er forbedret, hastigheden for RCC dødsfald fortsætter med at stige, hvilket antyder, at metastatisk sygdom er i høj grad ansvarlig for mortalitet 3. Terapi udvikling for RCC har udviklet i løbet af det sidste årti med opdagelsen og gennemførelsen af ​​målrettede behandlinger og immunoterapi. Desværre, progressionsfri survival og samlet overlevelse af avancerede tilfælde er stadig et godt stykke under 2 år for de fleste patienter 4, 5, 6. Disse statistikker garanterer behovet for yderligere forskning i identifikation og udvikling af effektive behandlinger for RCC. For i tilstrækkelig grad fremme terapeutiske indgreb for mRCC, der først behov translationelt-relevante prækliniske modeller af sygdommen. Udvikling af en relevant og konsekvent model til at rekapitulere den menneskelige tilstand af fremskreden RCC bør behandle flere vigtige spørgsmål: 1) er den model, anatomisk relevante; 2) Er tumoren fremskridt i lighed med den humane patologi; 3) Har metastaser opstår fra den primære tumor; og 4) Kan primær og metastatisk tumor progression overvåges over tid? Afhængigt af den type behandling, der undersøges, muse RCC tumorcellelinjer, humane RCC tumorcellelinjer og humant RCC patient-afledte xenogflåder kan anvendes i immunkompetente eller immundefekte mus. En vært med et intakt og funktionelt immunsystem er påkrævet for disse studier, der evaluerer nogle aspekter af immunterapi, der medfører anvendelse af den velbeskrevne Renca cellelinie afledt fra en spontan renal adenocarcinom Balb / c mus 7. De fleste undersøgelser injicere RENCA-celler subkutant (sc), som danner en let efter mål lokal tumor eller intravenøst (iv) i Balb / c-mus for at producere eksperimentelle lunge &quot;metastaser&quot; 8, 9, 10, 11, 12. Anvendelse af et sc-implanteret Renca tumor at modellere humant RCC har en række begrænsninger, herunder unøjagtig innervation af vaskulatur 13, forskelle i mikromiljø 14, 15, og en mangel på organer / tumor cellulære Communication 13, 16. Derudover har mange sc tumorer (især Renca) ikke metastaserer til distale organer, inhibering studiet af en begivenhed, som er en almindelig klinisk karakteristisk 17. At studere mRCC, kan RENCA-celler injiceres iv at etablere tumorbelastning i lungerne-primære placering metastaser i RCC patienter. Intravenøs injektion metode til at initiere metastatiske tumorer imidlertid ikke tillader undersøgelse af hvordan, hvornår eller hvorfor cellerne er migreret fra det primære organ (dvs. nyren) til fjernt sted. En model med både primær og metastatisk sygdom tydeligt i det samme dyr er afgørende for at studere progression og behandling af fremskreden sygdom, især i betragtning af at metastaser er typisk de årsag til dødelighed hos disse patienter. Vores laboratorium har udviklet en musemodel af mRCC, der inkorporerer alle de funktioner, der er skitseret ovenfor. At etablere PRimary, ortotope tumorer, er RENCA-celler implanteres direkte ind i nyren af ​​dyret gennem den gennemskinnelige peritoneum. Et lille snit i venstre flanke muliggør visualisering af milten (som en milepæl) og den venstre nyre. Anvendelse af en lille nål, er Renca tumorceller injiceres direkte i nyren gennem peritoneum for orthotopisk implantation. Sammenlignet med andre fremgangsmåder til at implantere RENCA-celler under nyrekapslen 18 denne form for implantation muliggør en højere kapacitet, som det er temmelig ikke-invasiv, kræver ikke suturering, er af en lav smerte klasse, og er tidsbesparende, når praktiseres. This velkarakteriserede model resulterer i reproducerbar primær tumorbelastning (~ 99% take rate) i det injicerede nyre samt metastatisk tumor belastning i lungerne. De væsentlige fordele ved denne model omfatter dens syngen natur, der giver mulighed for immunterapi undersøgelser; dens spontane metastaser, for at studere envanceret sygdom; og dens orthotopisk implantation, at modellere den anatomiske konsekvenser for sygdomsudvikling og behandling. Terapier rettet mod at målrette RCC vil få stor gavn af udnyttelsen af ​​denne model under præklinisk udvikling.

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			<td height="52" style="height:52px;width:88px;">Artificial tears ophthalmic ointment</td>
			<td style="width:88px;">Akorn</td>
			<td style="width:156px;">NDC 17478-162-35</td>
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			<td height="52" style="height:52px;width:88px;">Vetbond Tissue Adhesive</td>
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			<td height="69" style="height:69px;width:88px;">Betadine Solution (Povidone-iodine, 5%)</td>
			<td style="width:88px;">Purdue Products, L.P.</td>
			<td style="width:156px;">NDC 67618-155-32</td>
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			<td height="69" style="height:69px;width:88px;">0.25% Marcaine (bupivavaine HCL injection)</td>
			<td style="width:88px;">Hospira</td>
			<td style="width:156px;">0409-1587-50&#160;</td>
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			<td height="22" style="height:22px;width:88px;">Heating pad</td>
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			<td height="22" style="height:22px;width:88px;">Syringes</td>
			<td style="width:88px;">BD</td>
			<td style="width:156px;">309659</td>
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			<td height="22" style="height:22px;width:88px;">Gauze</td>
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			<td height="22" style="height:22px;width:88px;">Xylazine&#160;</td>
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			<td height="52" style="height:52px;width:88px;">Hank&#8217;s Balanced Salts</td>
			<td style="width:88px;">Sigma-Aldrich</td>
			<td style="width:156px;">H2387</td>
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			<td height="22" style="height:22px;width:88px;">IVIS</td>
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			<td height="52" style="height:52px;width:88px;">D-Luciferin, Potassium Salt</td>
			<td style="width:88px;">GoldBio</td>
			<td style="width:156px;">LUCK-1G</td>
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			<td height="22" style="height:22px;width:88px;">India Ink</td>
			<td style="width:88px;">Chartpak Inc</td>
			<td style="width:156px;">44201</td>
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			<td height="22" style="height:22px;width:88px;">Scissors</td>
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			<td height="22" style="height:22px;width:88px;">Forceps</td>
			<td style="width:88px;"></td>
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			<td height="69" style="height:69px;width:88px;">Sleeping Beauty (SB) transposon system</td>
			<td style="width:88px;">Neuromics</td>
			<td style="width:156px;">SBT0200</td>
			<td></td>
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			<td height="22" style="height:22px;width:88px;">Renca</td>
			<td style="width:88px;">ATCC</td>
			<td style="width:156px;">CRL-2947</td>
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a syngeneic mouse model of metastatic renal cell carcinoma for quantitative and longitudinal assessment of preclinical therapies

Renalcellecarcinom (RCC) påvirker&gt; 60.000 mennesker i USA hvert år, og ~ 30% af RCC-patienter har flere metastaser på tidspunktet for diagnosen. Metastatisk RCC (mRCC) er uhelbredelig, med en median overlevelsestid på kun 18 måneder. Immun-baserede interventioner (fx interferon (IFN) og interleukin (IL) -2) inducere varig respons på en brøkdel af MRCC patienter og multikinasehæmmer hæmmere (f.eks sunitinib eller sorafenib) eller anti-VEGF-receptor monoklonale antistoffer (mAb) er stort set palliativ, som komplette remissioner er sjældne. Sådanne mangler ved de nuværende behandlinger af MRCC patienter giver rationalet for udviklingen af ​​hidtil ukendte behandlingsprotokoller. En nøglekomponent i præklinisk afprøvning af nye terapier til mRCC er en egnet dyremodel. Gavnlige funktioner, der rekapitulere den menneskelige tilstand indbefatter en primær nyretumor, renale tumormetastaser, og et intakt immunsystem til at undersøge enhver terapi-drevet immun effector reaktioner og dannelsen af ​​tumor-induceret immunosuppressive faktorer. Denne rapport beskriver en ortotopisk mRCC musemodel, der har alle disse funktioner. Beskriver vi en intrarenal implantationsteknik hjælp af musen renal adenocarcinom cellelinie Renca, efterfulgt af vurdering af tumorvækst i nyren (primær sted) og lungerne (metastatisk site).

<html> Den vellykkede implantation af RENCA-celler vil resultere i tumorudvikling i nyren og metastase til lungerne af musene. Efter udskæring af tumorbærende og de kontralaterale nyrer blev våde væv vægte (g) målt for at påvise tumorbyrden baseret på forøget vægt (figur 1). RENCA-tumorer kan identificeres ved immunhistokemi gennem cytokeratin 8 og 18-farvning (figur 2). For langsgående undersøgelser blev en luciferase-udtrykkende Renca cellel...

Watch this Scientific Journal Video about A Syngeneic Mouse Model of Metastatic Renal Cell Carcinoma for Quantitative and Longitudinal Assessment of Preclinical Therapies at JoVE.com

A Syngeneic Mouse Model of Metastatic Renal Cell Carcinoma for Quantitative and Longitudinal Assessment of Preclinical Therapies

Gennemførelse af en orthotopisk model af renalcellekarcinom i immunokompetente mus giver investigator en klinisk relevant system, defineret ved tilstedeværelsen af ​​en primær nyretumor og lungemetastaser i det samme dyr. Dette system kan anvendes til præklinisk afprøve forskellige behandlinger in vivo.

En syngen musemodel af metastaserende renalcellecarcinom for Kvantitativ og Longitudinal Vurdering af Prækliniske Terapier

Renal cell carcinoma (RCC) affects &#62; 60,000 people in the United States annually, and ~ 30% of RCC patients have multiple metastases at the time of ...

Research

JoVE Journal

Cancer Research

Development and Maintenance of a Preclinical Patient Derived Tumor Xenograft Model for the Investigation of Novel Anti-Cancer Therapies

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Patient derived tumor xenograft (PDTX) models provide a necessary platform in facilitating anti-cancer drug development prior to human trials. Human tumor ...

Surgical Procedures and Methodology for a Preclinical Murine Model of De Novo Mammary Cancer Metastasis

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Cancer cachexia is the progressive loss of skeletal muscle mass and adipose tissue, negative nitrogen balance, anorexia, fatigue, inflammation, and ...

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Perineural invasion (PNI) is found in approximately 40% of head and neck squamous cell carcinomas (HNSCC). Despite multimodal treatment with surgery, ...

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Current treatment options for advanced and recurrent head and neck squamous cell carcinoma (HNSCC) enclose radiation and chemo-radiation approaches with ...

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En Syngeneic bugspytkirtelkræft musen Model til at studere virkningerne af irreversibel elektroporation

Pancreatic cancer (PC), a disease which kills approximately 40,000 patients each year in the US, has successfully evaded several therapeutic approaches ...

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Orthotopic tumor modeling is a valuable tool for pre-clinical prostate cancer research, as it has multiple advantages over both subcutaneous and ...

A Syngeneic Mouse B-Cell Lymphoma Model for Pre-Clinical Evaluation of CD19 CAR T Cells

Syngeneic musen B-celle lymfom Model for præ-klinisk evaluering af CD19 bil T celler

The astonishing clinical success of CD19 chimeric antigen receptor (CAR) T-cell therapy has led to the approval of two second generation chimeric antigen ...

Intramucosal Inoculation of Squamous Cell Carcinoma Cells in Mice for Tumor Immune Profiling and Treatment Response Assessment

Intramucosal podning af pladecellekræft celler i mus til Tumor immun profilering og behandling svar vurdering

Head and neck squamous cell carcinoma (HNSCC) is a debilitating and deadly disease with a high prevalence of recurrence and treatment failure. To develop ...

Micromanipulation of Circulating Tumor Cells for Downstream Molecular Analysis and Metastatic Potential Assessment

Micromanipulation af cirkulerende tumor celler til downstream Molekylær analyse og metastatisk potentiel vurdering

Blood-borne metastasis accounts for&#160;most cancer-related deaths and involves circulating tumor cells (CTCs) that are successful in establishing new ...