Utilize este identificador para referenciar este registo: http://hdl.handle.net/10451/9695
Título: Re and 99m Tc tricarbonyl probes for target-specific detection of melanoma and sentinel lymph node
Autor: Morais, Maurício da Silva, 1978-
Orientador: Correia, João Domingos Galamba, 1967-
Santos, Isabel Rego dos, 1948-
Palavras-chave: Melanoma
Nódulo linfático
Imagiologia
Tomografia por emissão de fotão único (SPECT)
Teses de doutoramento - 2013
Data de Defesa: 2013
Resumo: The work described in this thesis aimed at the development of M(I) (M = 99mTc, Re) specific probes for the detection of malignant melanoma and sentinel lymph node (SLN) through the in vivo targeting of membrane receptors. In the former case we have designed M(CO)3-complexes stabilized by tridentate chelators containing a pyrazolyl-diamine chelating unit (N,N,N donor atom set) and pendant α-melanocyte-stimulating hormone (α-MSH) derivatives for targeting the melanocortin receptor 1 (MC1R), which is overexpressed in melanotic and amelanotic human melanoma cells. Among the various strategies described herein to improve both the MC1R targeting properties and the biological profile of 99mTc-labeled α-MSH analogs, the chapter 2 of this thesis focus on the design of homobivalent constructs containing two copies of a linear α-MSH analog (NAPamide) separated by linkers of different nature (L5, symmetric alkyl chain; L6, asymmetric semirigid spacer) and length (L5, 9 atoms; L6, 14 atoms). The MC1R-binding affinity of both peptide conjugates is significantly higher than that of the monovalent conjugate L4. Metallation of L4 - L6 yielded complexes of the type fac-[M(CO)3(κ3-L)]+ (M = 99mTc/Re; Tc4/Re4, L = L4; Tc5/Re5, L = L5; Tc6/Re6, L = L6), with the rhenium compounds presenting IC50 values in the sub- and nanomolar range, which are still better than or comparable to the IC50 values of the non-conjugated NAPamide. The MC1R-mediated internalization of Tc5 and Tc6 in B16F1 melanoma cells is higher than that of Tc4. However, biodistribution studies in melanoma-bearing mice have shown low tumor uptake and significant accumulation of radioactivity in kidneys. Moreover, no correlation between tumor uptake and valency was found. Chapter 3 describes novel peptide conjugates (L7 and L8), which contain a thioether (SNBA-MSHhex) or amine (NNBA-MSHhex) bridge-cyclized α-MSH analog, respectively, and a pyrazolyl-diamine chelating unit. Metallation of L7 and L8 gave complexes of the type fac-[M(CO)3(κ3-L)]+ (M = 99mTc/Re; Tc7/Re7, L = L7; Tc8/Re8, L = L8). Competitive binding affinity assays demonstrated that the compounds containing the alkylamine bridge-cyclized peptide moiety present lower IC50 values than the one cyclized via the alkylthioaryl bridge. NMR structural analysis suggested that such difference could be ascribed to an anisotropic effect arising from the interaction between the Arg side chain and the surrounding aromatic residues in the pharmacophore sequence (His-DPhe-Arg-Trp) of SNBA-MSHhex, hampering a favorable interaction of the peptide with the MC1R. NMR studies have also shown that the three-dimensional structural features of the peptide moiety were conserved upon conjugation to the chelator and, most importantly, after metallation. The chapter 4 of this thesis reports the impact of pyrazolyl-diamine chelators with different azolyl-ring substitution patterns (carboxylate at the 4-position and methyl groups at the 3,5 positions, L3; no substituent groups, L10; carboxylate at the 4-position, L11) on the MC1R-targeting properties and pharmacokinetic profile of a 99mTc(CO)3-labeled lactam bridge-cyclized α-MSH analog (βAlaNleCycMSHhex). Conjugation of those bifunctional chelators to βAlaNleCycMSHhex yielded the peptide conjugates L13 – L15, which upon reaction with fac-[99mTc(CO)3(H2O)3]+ gave the (radio)metallated compounds Tc13 – Tc15. Biodistribution studies in murine melanoma-bearing mice have shown that all radiopeptides (Tc13 – Tc15) presented a good melanoma uptake (≥ 9.90  1.10 % IA/g). The introduction of a carboxylate group in the azolyl-ring lead to a remarkable reduction of kidney (> 89 %) and liver (> 91 %) uptake for Tc14 and Tc15, respectively, when compared to the lead radiopeptide Tc9, which has methyl groups at the 3,5 positions of the azolyl-ring. The remarkable tumor uptake and favorable tumor/non-target organs ratios of Tc14 and Tc15 highlights the potential of both compounds as melanoma imaging agents. Chapter 5 reports the design, characterization and biological evaluation of mannosylated dextran derivatives with pyrazolyl-diamine chelating units for sentinel lymph node detection (SLND) through mannose receptor (MR) targeting. The mannosylated dextran derivatives L18 and L19 contain, respectively, 4 and 8 chelating units per mol of dextran to stabilize the fac-[M(CO)3]+ moiety (M = Re, 99mTc), as well as 13 mannose units per mol of dextran for MR targeting. A bimodal probe for SLND by SPECT and optical imaging in the near infrared (NIR) field is also reported in chapter 5. L20 was prepared by conjugation of 3 to 4 fluorophore units to L19. Metallation of L18 – L20 gave polymeric complexes of the type fac-[M(CO)3(κ3-L)]+ (M = 99mTc/Re; Tc18/Re18, L = L18; Tc19/Re19, L = L19; Tc20/Re20, L = L20). Physical characterization of the mannosylated dextran derivatives has shown that the hydrodynamic diameter and the zeta potencial of the nanoparticles were affected by chemical modification. Indeed, the hydrodynamic diameter of L18 – L20 are within the range of 7.0 to 17.7 nm while the diameter of the respective metallated compounds (Re18 – Re20) is slightly higher, within the range 8.4 to 22.5 nm. Biological evaluation of the polymeric radiotracers in Wistar rats has shown a good accumulation in the popliteal (sentinel) node at 1 h p.i. (6.71 ± 2.35 % IA/organ,structural features of the peptide moiety were conserved upon conjugation to the chelator and, most importantly, after metallation. The chapter 4 of this thesis reports the impact of pyrazolyl-diamine chelators with different azolyl-ring substitution patterns (carboxylate at the 4-position and methyl groups at the 3,5 positions, L3; no substituent groups, L10; carboxylate at the 4-position, L11) on the MC1R-targeting properties and pharmacokinetic profile of a 99mTc(CO)3-labeled lactam bridge-cyclized α-MSH analog (βAlaNleCycMSHhex). Conjugation of those bifunctional chelators to βAlaNleCycMSHhex yielded the peptide conjugates L13 – L15, which upon reaction with fac-[99mTc(CO)3(H2O)3]+ gave the (radio)metallated compounds Tc13 – Tc15. Biodistribution studies in murine melanoma-bearing mice have shown that all radiopeptides (Tc13 – Tc15) presented a good melanoma uptake (≥ 9.90  1.10 % IA/g). The introduction of a carboxylate group in the azolyl-ring lead to a remarkable reduction of kidney (> 89 %) and liver (> 91 %) uptake for Tc14 and Tc15, respectively, when compared to the lead radiopeptide Tc9, which has methyl groups at the 3,5 positions of the azolyl-ring. The remarkable tumor uptake and favorable tumor/non-target organs ratios of Tc14 and Tc15 highlights the potential of both compounds as melanoma imaging agents. Chapter 5 reports the design, characterization and biological evaluation of mannosylated dextran derivatives with pyrazolyl-diamine chelating units for sentinel lymph node detection (SLND) through mannose receptor (MR) targeting. The mannosylated dextran derivatives L18 and L19 contain, respectively, 4 and 8 chelating units per mol of dextran to stabilize the fac-[M(CO)3]+ moiety (M = Re, 99mTc), as well as 13 mannose units per mol of dextran for MR targeting. A bimodal probe for SLND by SPECT and optical imaging in the near infrared (NIR) field is also reported in chapter 5. L20 was prepared by conjugation of 3 to 4 fluorophore units to L19. Metallation of L18 – L20 gave polymeric complexes of the type fac-[M(CO)3(κ3-L)]+ (M = 99mTc/Re; Tc18/Re18, L = L18; Tc19/Re19, L = L19; Tc20/Re20, L = L20). Physical characterization of the mannosylated dextran derivatives has shown that the hydrodynamic diameter and the zeta potencial of the nanoparticles were affected by chemical modification. Indeed, the hydrodynamic diameter of L18 – L20 are within the range of 7.0 to 17.7 nm while the diameter of the respective metallated compounds (Re18 – Re20) is slightly higher, within the range 8.4 to 22.5 nm. Biological evaluation of the polymeric radiotracers in Wistar rats has shown a good accumulation in the popliteal (sentinel) node at 1 h p.i. (6.71 ± 2.35 % IA/organ,xiii Tc18; and 7.53 ± 0.69 % IA/organ, Tc19), with significant retention up to 3 h, which is mediated by MR. The high popliteal extraction value of Tc19 (94.47 ± 2.45 % at 1 h p.i.) highlights its potential as a very promising radiotracer to be further explored as SLN imaging agent in humans. The biological evalution of Tc20 has shown that the incorporation of a fluorophore into the nanoparticle did not not alter the pharmacokinetic properties of the bimodal probe as compared to the radiotracer Tc19. In fact, Tc20 and Tc19 presented high popliteal extraction value (89.66 ± 0.03 % and 87.81 ± 3.75 %, respectively) at 3 h p.i. Furthermore, the biological evaluation of Tc20 demonstrated a perfect coregistration of the two signals, allowing nuclear and optical imaging techniques to complement each other in the detection and excision of the popliteal lymph node.
Descrição: Tese de doutoramento, Química (Química Inorgânica), Universidade de Lisboa, Faculdade de Ciências, 2013
URI: http://hdl.handle.net/10451/9695
Aparece nas colecções:FC - Teses de Doutoramento

Ficheiros deste registo:
Ficheiro Descrição TamanhoFormato 
ulsd066946_td_Mauricio_Morais.pdf8,66 MBAdobe PDFVer/Abrir


FacebookTwitterDeliciousLinkedInDiggGoogle BookmarksMySpace
Formato BibTex MendeleyEndnote Degois 

Todos os registos no repositório estão protegidos por leis de copyright, com todos os direitos reservados.