Modeling interstitial fluid flow involves processes such as fluid diffusion, convective transport in the extracellular matrix, and extravasation from blood vessels. In all of these processes, computational fluid dynamics can play a crucial role in elucidating the mechanisms of fluid flow in solid tumors and surrounding tissues. To date, microvasculature flow modeling has been most extensively studied with simple tumor shapes and their capillaries at different levels and scales. With our proposed numerical model, however, more complex and realistic tumor shapes and capillary networks can be studied. First, a mathematical model of interstitial fluid flow is developed, based on the application of the governing equations for fluid flow, i.e., the conservation laws for mass and momentum, to physiological systems containing solid tumors. Simulations of interstitial fluid transport in a homogeneous solid tumor demonstrate that, in a uniformly perfused tumor, i.e., one with no necrotic region, the interstitial pressure distribution results in a non-uniform distribution of drug particles. Pressure distribution for different values of necrotic radii is examined, and two new parameters, the critical tumor radius and critical necrotic radius, are defined. In specific ranges of these critical dimensions the interstitial fluid pressure is relatively lower, which in turn leads to a diminished opposing force against drug movement and a subsequently higher drug concentration and potentially enhanced therapeutic effects. In this work, the numerical model of fluid flow in solid tumors is further developed to incorporate and investigate non-spherical tumor shapes such as prolate and oblate ones. Using this enhanced model, tumor shape and size effects on drug delivery to solid tumors are then studied. Based on the assumption that drug particles flow with the interstitial fluid, the pressure and velocity maps of the latter are used to illustrate the drug delivery pattern in a solid tumor. Additionally, the effects of the surface area per unit volume of the tissue, as well as vascular and interstitial hydraulic conductivity on drug delivery efficiency, are investigated. Using a tumor-induced microvasculature architecture instead of a uniform distribution of vessels provides a more realistic model of solid tumors. To this end, continuous and discrete mathematical models of angiogenesis were utilized to observe the effect of matrix density and matrix degrading enzymes on capillary network formation in solid tumors. Additionally, the interactions between matrix-degrading enzymes, the extracellular matrix and endothelial cells are mathematically modeled. Existing continuous and discrete models of angiogenesis were modified to impose the effect of matrix density on the solution. The imposition has been performed by a specific function in movement potential. Implementing realistic boundary and initial conditions showed that, unlike in previous models, the endothelial cells accelerate as they migrate toward the tumor. Now, the tumor-induced microvasculature network can be applied to the model developed in Chapters 2 and 3. Once the capillary network was set up, fluid flow in normal and cancerous tissues was numerically simulated under three conditions: constant and uniform distribution of intravascular pressure in the whole domain, a rigid vascular network, and an adaptable vascular network. First, governing equations of sprouting angiogenesis were implemented to specify the different domains for the network and interstitium. Governing equations for flow modeling were introduced for different domains. The conservation laws for mass and momentum, Darcy’s equation for tissue, and a simplified Navier Stokes equation for blood flow through capillaries were then used for simulating interstitial and intravascular flows. Finally, Starling’s law was used to close this system of equations and to couple the intravascular and extravascular flows. The non-continuous behavior of blood and the adaptability of capillary diameter to hemodynamics and metabolic stimuli were considered in blood flow simulations through a capillary network. This approach provided a more realistic capillary distribution network, very similar to that of the human body. This work describes the first study of flow modeling in solid tumors to realistically couple intravascular and extravascular flow through a network generated by sprouting angiogenesis, consisting of one parent vessel connected to the network. Other key factors incorporated in the model for the first time include capillary adaptation, non-continuous viscosity blood, and phase separation of blood flow in capillary bifurcation. Contrary to earlier studies which arbitrarily assumed veins and arteries to operate on opposite sides of a tumor network, the present approach requires the same vessel to run and from the network. Expanding the earlier models by introducing the outlined components was performed in order to achieve a more-realistic picture of blood flow through solid tumors. Results predict an almost doubled interstitial pressure and are in better agreement with human biology compared to the more simplified models generally in use today.

Modeling interstitial fluid flow involves processes such as fluid diffusion, convective transport in the extracellular matrix, and extravasation from blood vessels. In all of these processes, computational fluid dynamics can play a crucial role in elucidating the mechanisms of fluid flow in solid tumors and surrounding tissues. To date, microvasculature flow modeling has been most extensively studied with simple tumor shapes and their capillaries at different levels and scales. With our proposed numerical model, however, more complex and realistic tumor shapes and capillary networks can be studied. First, a mathematical model of interstitial fluid flow is developed, based on the application of the governing equations for fluid flow, i.e., the conservation laws for mass and momentum, to physiological systems containing solid tumors. Simulations of interstitial fluid transport in a homogeneous solid tumor demonstrate that, in a uniformly perfused tumor, i.e., one with no necrotic region, the interstitial pressure distribution results in a non-uniform distribution of drug particles. Pressure distribution for different values of necrotic radii is examined, and two new parameters, the critical tumor radius and critical necrotic radius, are defined. In specific ranges of these critical dimensions the interstitial fluid pressure is relatively lower, which in turn leads to a diminished opposing force against drug movement and a subsequently higher drug concentration and potentially enhanced therapeutic effects. In this work, the numerical model of fluid flow in solid tumors is further developed to incorporate and investigate non-spherical tumor shapes such as prolate and oblate ones. Using this enhanced model, tumor shape and size effects on drug delivery to solid tumors are then studied. Based on the assumption that drug particles flow with the interstitial fluid, the pressure and velocity maps of the latter are used to illustrate the drug delivery pattern in a solid tumor. Additionally, the effects of the surface area per unit volume of the tissue, as well as vascular and interstitial hydraulic conductivity on drug delivery efficiency, are investigated. Using a tumor-induced microvasculature architecture instead of a uniform distribution of vessels provides a more realistic model of solid tumors. To this end, continuous and discrete mathematical models of angiogenesis were utilized to observe the effect of matrix density and matrix degrading enzymes on capillary network formation in solid tumors. Additionally, the interactions between matrix-degrading enzymes, the extracellular matrix and endothelial cells are mathematically modeled. Existing continuous and discrete models of angiogenesis were modified to impose the effect of matrix density on the solution. The imposition has been performed by a specific function in movement potential. Implementing realistic boundary and initial conditions showed that, unlike in previous models, the endothelial cells accelerate as they migrate toward the tumor. Now, the tumor-induced microvasculature network can be applied to the model developed in Chapters 2 and 3. Once the capillary network was set up, fluid flow in normal and cancerous tissues was numerically simulated under three conditions: constant and uniform distribution of intravascular pressure in the whole domain, a rigid vascular network, and an adaptable vascular network. First, governing equations of sprouting angiogenesis were implemented to specify the different domains for the network and interstitium. Governing equations for flow modeling were introduced for different domains. The conservation laws for mass and momentum, Darcy’s equation for tissue, and a simplified Navier Stokes equation for blood flow through capillaries were then used for simulating interstitial and intravascular flows. Finally, Starling’s law was used to close this system of equations and to couple the intravascular and extravascular flows. The non-continuous behavior of blood and the adaptability of capillary diameter to hemodynamics and metabolic stimuli were considered in blood flow simulations through a capillary network. This approach provided a more realistic capillary distribution network, very similar to that of the human body. This work describes the first study of flow modeling in solid tumors to realistically couple intravascular and extravascular flow through a network generated by sprouting angiogenesis, consisting of one parent vessel connected to the network. Other key factors incorporated in the model for the first time include capillary adaptation, non-continuous viscosity blood, and phase separation of blood flow in capillary bifurcation. Contrary to earlier studies which arbitrarily assumed veins and arteries to operate on opposite sides of a tumor network, the present approach requires the same vessel to run and from the network. Expanding the earlier models by introducing the outlined components was performed in order to achieve a more-realistic picture of blood flow through solid tumors. Results predict an almost doubled interstitial pressure and are in better agreement with human biology compared to the more simplified models generally in use today.

In the past several years, cancer treatments using FDA approved immune checkpoint inhibitors have become more popular than common chemotherapeutic agents. However, the costs, risks, and benefits associated with these treatments are still being assessed. Currently, new therapies are being tested that utilize oncolytic viruses to treat solid tumor and metastatic cancers. Reovirus is a non-enveloped virus with a double capsid structure and a genome consisting of 10 segments of double-stranded RNA encoding eleven proteins, which has been shown to have effective oncolytic activity. There are various different strains of reoviruses that can produce cytopathic effects in mammalian host cells. Moreover, several studies have shown that reovirus can be administered in multiple ways and that administration may depend on the cancer type. This investigation examined the type 3 dearing strain of reovirus and whether different types of tumors would benefit from having a specific administration appropriate to their type such as intratumoral injections for solid tumors and intravenous administration for blood/metastatic cancers. Various clinical trials were assessed in which reovirus was administered intratumorally, intravenously at a maximum dose of 3x1010 TCID50, and in combinations with other cancer therapeutics. Reovirus was shown to be safe and well-tolerated across a variety of administrations and cancer morphologies. Moreover, along with its cytopathic effects, reovirus was shown to have potent immune system stimulating effects. Overall, intratumoral administration was preferred effective for solid tumor cancers while intravenous administration was preferred for blood and metastatic cancers.

No description available.

Un nombre croissant d'observations suggèrent que les pDC sont fortement impliqués dans le cancer, car les pDC sont recrutés dans des tumeurs solides, tant chez les patients humains que chez les modèles murins et sont généralement en corrélation avec un mauvais pronostique. Les pDC infiltrant les tumeurs présentent souvent un phénotype immature et sont des pauvres producteurs de IFN-I, de cytokines et de chimiokines pro-inflammatoires en réponse à la stimulation TLR, contribuant à l'établissement d'un micro-environnement tumoral immunosuppresseur qui favorise la croissance tumorale. D'autre part, lorsqu'ils sont activés à l'intérieur de la tumeur, les pDC pourraient être capable de générer des réponses immunitaires antitumorales efficaces qui contribueraient à la régression tumorale. Ce double rôle de TA-pDC rend cette population cellulaire d'un grand intérêt pour être ciblée dans l'immunothérapie tumorale. Par conséquent, notre étude est centrée sur l'exploration du rôle de pDC dans l'orchestration des réponses immunitaires antitumorales, visant à déchiffrer les caractéristiques fonctionnelles de pDC dans le micro-environnement de la tumeur.Nos résultats montrent que les pDC sont recrutés dans le micro-environnement de la tumeur TC-1 et B16-OVA, et les pDC infiltrant les tumeurs dans les deux modèles de tumeurs présentent un profil d'activation et d'expression génique distinct par rapport aux pDC purifiés à partir de la rate naïve, ce qui suggère un effet du microenvironnement tumoral dans le phénotype et les fonctions de TC-1 et BDC-OVA infiltrant pDC. En fait, les facteurs solubles sécrétés par les cellules tumorales TC-1 et B16-OVA et présentes dans le micro-environnement de la tumeur sont capables d'affecter fortement les fonctions des pDC, notamment leur capacité à produire IFN-α après la stimulation TLR-9. Parmi les facteurs solubles présents dans le micro-environnement de la tumeur, nos résultats montrent que le TGF-β seul est capable de bloquer la production d'IFN-α, suggérant clairement une influence du TGF-β dans les mécanismes intracellulaires conduisant à la production d'IFN-I par pDC. En outre, notre étude révèle que les effets du TGF-β sur le pDC affectent aussi les capacités de production de l'IFN-I, mais aussi leur capacité à sécréter d'autres cytokines et chimiokines ainsi que leur phénotype. Enfin, l'étude de l'environnement tumoral TC-1 en l'absence de pDC démontre un rôle délétère de cette population dans la croissance de la tumeur et montre une fonction claire de pDC dans l'induction de réponses immunitaires anti-tumorales dépendantes de NK.Les résultats obtenus au cours de cette thèse de doctorat ont mis en évidence le rôle important de la PDC dans le contexte tumoral et ont permis une meilleure compréhension du ciblage de cette population cellulaire contre l'immunothérapie tumorale / A growing number of observations suggest that pDC are highly implicated in cancer, since pDC are recruited into solid tumors, both in human patients and in murine models and generally correlate with a poor clinical outcome. Tumor infiltrating pDC often exhibit an immature phenotype and are poor producers of IFN-I and pro-inflammatory cytokines and chemokines in response to TLR stimulation, contributing to the establishment of an immunosuppressive tumor microenvironment that promotes tumor growth. On the other hand, when activated inside the tumor, pDC could be able to generate efficient anti-tumor immune responses that would contribute to tumor regression. This dual role of TA-pDC renders this cell population of great interest to be targeted in tumor immunotherapy. Hence, our study is centered in exploring the role of pDC in the orchestration of anti-tumor immune responses, aiming to decipher functional characteristics of pDC within the tumor microenvironment. Our results show that pDC are recruited into the TC-1 and B16-OVA tumor microenvironment and tumor infiltrating pDC in both tumor models exhibit a distinct activation and gene expression profile as compared to pDC purified from naïve spleen, suggesting an effect of the tumor microenvironment in the phenotype and functions of TC-1 and B16-OVA infiltrating pDC. In fact, the soluble factors secreted by the TC-1 and B16-OVA tumor cells and present in the tumor microenvironment are able to greatly affect pDC functions, more importantly their ability to produce IFN-α following TLR-9 stimulation. Among the soluble factors present in the tumor microenvironment, our results show that TGF-β alone is able to impair the production of IFN-α, clearly suggesting an influence of TGF-β in the intracellular machinery leading to IFN-I production by pDC. In addition, our study reveals that the effects of TGF-β on pDC are broader affecting not only the IFN-I producing abilities, but also their capacity to secrete other cytokines and chemokines as well as their phenotype. Finally, the study of the TC-1 tumor environment in the absence of pDC demonstrates a deleterious role of this population in the tumor growth and shows a clear function of pDC in the induction of NK-dependent anti-tumor immune responses.The results obtained throughout this PhD thesis highlighted the important role of pDC in the tumoral context and allowed further insight in targeting this cell population to tumor immunotherapy

Autoimmune hemolytic anemia (AIHA) is a disease process that involves the destruction of red blood cells mediated by the humoral immune system. It can be characterized as a cold agglutinin syndrome, paroxysmal cold hemoglobinuria, and warm, mixed type, and drug-induced AIHA. Although a well-established relationship exists between the presence of AIHA and lymphoproliferative malignancy, AIHA rarely presents in association with solid malignancies. An analysis of the limited number of published cases of AIHA in association with solid malignancies performed showed that AIHA may present before the diagnosis of a solid malignancy, concurrently with the presence of a solid malignancy, or even on resolution of a solid malignancy. Few cases of solid cancers associated with AIHA have been reported. AIHA rarely presents as a paraneoplastic syndrome indicating existence of a solid cancer. We report a case of inflammatory breast cancer with AIHA.

A terapia fotodinâmica (Photodynamic Therapy - PDT) é um método de tratar neoplasias baseado na interação entre luz, oxigênio molecular e um agente fotossensibilizador. Após a administração do agente, o tumor é iluminado com luz visível, ativando-o e produzindo espécies reativas de oxigênio, altamente citotóxicas, que provocam morte celular e destruição tecidual. Com a destruição do tumor há ativação do sistema imune inato e o subsequente processo inflamatório determina a apresentação de antígenos tumorais aos linfócitos, promovendo uma resposta imunológica adaptativa contra o tecido tumoral. O presente trabalho visou estudar a PDT associando um laser de diodo como fonte de luz e o fotossensibilizante Azul de Metileno (AM) a 1%, avaliando a sua efetividade no tratamento do Tumor de Ehrlich (TE) em sua forma sólida e a resposta imunológica nos animais tratados. Em um primeiro estudo, avaliou-se macro e microscopicamente tumores tratados, determinando a capacidade do protocolo em induzir inflamação e destruição do tecido tumoral. No segundo estudo, a resposta imune foi estudada em camundongos desafiados com um segundo implante de células do tumor de Ehrlich. O primeiro implante tumoral foi tratado com a PDT ou a excisão cirúrgica, comparando-se com um grupo controle sem tratamento. Os parâmetros avaliados após 17 dias foram o crescimento tumoral (p>0,05), peso relativo dos órgãos linfóides [Baço (p<0,05) e Linfonodo poplíteo (p>0,05)], tamanho relativo do linfonodo (p<0,05), presença de metástase em linfonodo poplíteo (p>0,05), contagem de leucócitos sanguíneos (p>0,05) e análise morfométrica quantitativa do tumor secundário [determinação da fração volumétrica de células tumorais (p<0,05), infiltrado inflamatório (p<0,05), necrose (p>0,06) e porcentagem da área tumoral em necrose (p<0,05)]. A PDT com o AM foi capaz de induzir necrose do TE e inflamação, havendo diferenças da resposta imune sistêmica quando comparado aos animais tratados por meio de excisão cirúrgica do tumor de Ehrlich. / Photodynamic therapy (PDT) is a method of treating neoplasms based on the interaction between light, molecular oxygen and a photosensitizing agent. After administration of the photosensitizer, the tumor is illuminated with visible light, activating the agent and producing reactive oxygen species (ROS). This highly cytotoxic ROS cause cell death and tissue destruction. The activation of the innate immune system and the subsequent inflammation induces tumor antigen presentation to lymphocytes, promoting an adaptive immune response against the tumor cells. This work aimed to study the PDT using a diode laser as light source and Methylene Blue (MB) 1% as photosensitizer. It was accessed its effectiveness in treating Ehrlich Solid tumor (ET) and the immune response produced in treated animals. First the treated tumors were evaluated macroscopically and microscopically, determining the ability of the protocol to induce inflammation and tumor tissue destruction. In a second study, the immune response was studied in mice challenged with a second tumor cell implant. The primary tumor was treated with PDT or surgical excision, comparing with a control group without treatment. The parameters evaluated after 17 days were tumor growth (p> 0.05), relative weight of lymphoid organs [spleen (p <0.05) and popliteal lymph node (p> 0.05)], the relative size of the lymph node (p <0, 05), metastasis at lymph node (p>0,05), blood leukocyte count (p> 0.05) and quantitative morphometric analysis of secondary tumor [determining the volume fraction of tumor cells (p <0.05), inflammatory infiltrate (p <0.05), necrosis (p> 0.06) and tumor necrosis area (p <0.05)]. PDT with MB was able to induce necrosis of the ET and inflammation, with differences in the immune response when compared to animals treated surgically to remove the Ehrlich tumor in its solid form.

A terapia fotodinâmica (Photodynamic Therapy - PDT) é um método de tratar neoplasias baseado na interação entre luz, oxigênio molecular e um agente fotossensibilizador. Após a administração do agente, o tumor é iluminado com luz visível, ativando-o e produzindo espécies reativas de oxigênio, altamente citotóxicas, que provocam morte celular e destruição tecidual. Com a destruição do tumor há ativação do sistema imune inato e o subsequente processo inflamatório determina a apresentação de antígenos tumorais aos linfócitos, promovendo uma resposta imunológica adaptativa contra o tecido tumoral. O presente trabalho visou estudar a PDT associando um laser de diodo como fonte de luz e o fotossensibilizante Azul de Metileno (AM) a 1%, avaliando a sua efetividade no tratamento do Tumor de Ehrlich (TE) em sua forma sólida e a resposta imunológica nos animais tratados. Em um primeiro estudo, avaliou-se macro e microscopicamente tumores tratados, determinando a capacidade do protocolo em induzir inflamação e destruição do tecido tumoral. No segundo estudo, a resposta imune foi estudada em camundongos desafiados com um segundo implante de células do tumor de Ehrlich. O primeiro implante tumoral foi tratado com a PDT ou a excisão cirúrgica, comparando-se com um grupo controle sem tratamento. Os parâmetros avaliados após 17 dias foram o crescimento tumoral (p>0,05), peso relativo dos órgãos linfóides [Baço (p<0,05) e Linfonodo poplíteo (p>0,05)], tamanho relativo do linfonodo (p<0,05), presença de metástase em linfonodo poplíteo (p>0,05), contagem de leucócitos sanguíneos (p>0,05) e análise morfométrica quantitativa do tumor secundário [determinação da fração volumétrica de células tumorais (p<0,05), infiltrado inflamatório (p<0,05), necrose (p>0,06) e porcentagem da área tumoral em necrose (p<0,05)]. A PDT com o AM foi capaz de induzir necrose do TE e inflamação, havendo diferenças da resposta imune sistêmica quando comparado aos animais tratados por meio de excisão cirúrgica do tumor de Ehrlich. / Photodynamic therapy (PDT) is a method of treating neoplasms based on the interaction between light, molecular oxygen and a photosensitizing agent. After administration of the photosensitizer, the tumor is illuminated with visible light, activating the agent and producing reactive oxygen species (ROS). This highly cytotoxic ROS cause cell death and tissue destruction. The activation of the innate immune system and the subsequent inflammation induces tumor antigen presentation to lymphocytes, promoting an adaptive immune response against the tumor cells. This work aimed to study the PDT using a diode laser as light source and Methylene Blue (MB) 1% as photosensitizer. It was accessed its effectiveness in treating Ehrlich Solid tumor (ET) and the immune response produced in treated animals. First the treated tumors were evaluated macroscopically and microscopically, determining the ability of the protocol to induce inflammation and tumor tissue destruction. In a second study, the immune response was studied in mice challenged with a second tumor cell implant. The primary tumor was treated with PDT or surgical excision, comparing with a control group without treatment. The parameters evaluated after 17 days were tumor growth (p> 0.05), relative weight of lymphoid organs [spleen (p <0.05) and popliteal lymph node (p> 0.05)], the relative size of the lymph node (p <0, 05), metastasis at lymph node (p>0,05), blood leukocyte count (p> 0.05) and quantitative morphometric analysis of secondary tumor [determining the volume fraction of tumor cells (p <0.05), inflammatory infiltrate (p <0.05), necrosis (p> 0.06) and tumor necrosis area (p <0.05)]. PDT with MB was able to induce necrosis of the ET and inflammation, with differences in the immune response when compared to animals treated surgically to remove the Ehrlich tumor in its solid form.

In this Thesis, we have developed a microtechnology-based method for culturing and visualizing high numbers of individual cells and cell-cell interactions over extended periods of time. The foundation of the device is a silicon-glass multiwell microplate (also referred as microchip) directly compatible with fluorescence microscopy. The initial microchip design involved thousands of square wells of sizes up to 80 µm, for screening large numbers of cell-cell interactions at the single cell level. Biocompatibility and confinement tests proved the feasibility of the idea, and further investigation showed the conservation of immune cellular processes within the wells. Although the system is very reliable for screening, limitations related to synchronization of the interaction events, and the inability to maintain conjugations for long time periods, led to the development of a novel ultrasonic manipulation multiwell microdevice. The main components of the ultrasonic device is a 100-well silicon-glass microchip and an ultrasonic transducer. The transducer is used for ultrasonic actuation on the chip with a frequency causing half-wave resonances in each of the wells (2.0-2.5 MHz for wells with sizes 300-350 µm). Therefore, cells in suspension are directed by acoustic radiation forces towards a pressure node formed in the center of each well. This method allows simultaneous aggregation of cells in all wells and sustains cells confined within a small area for long time periods (even up to several days). The biological target of investigation in this Thesis is the natural killer (NK) cells and their functional properties. NK cells belong to the lymphatic group and they are important factors for host defense and immune regulation. They are characterized by the ability to interact with virus infected cells and cancer cells upon contact, and under suitable conditions they can induce target cell death. We have utilized the ultrasonic microdevice to induce NK-target cell interactions at the single cell level. Our results confirm a heterogeneity within IL-2 activated NK cell populations, with some cells being inactive, while others are capable to kill quickly and in a consecutive manner. Furthermore, we have integrated the ultrasonic microdevice in a temperature regulation system that allows to actuate with high-voltage ultrasound, but still sustain the cell physiological temperature. Using this system we have been able to induce formation of up to 100 solid tumors (HepG2 cells) in parallel without using surface modification or hydrogels. Finally, we used the tumors as targets for investigating NK cells ability to infiltrate and kill solid tumors.  To summarize, a method is presented for investigating individual NK cell behavior against target cells and solid tumors. Although we have utilized our technique to investigate NK cells, there is no limitation of the target of investigation. In the future, the device could be used for any type of cells where interactions at the single cell level can reveal critical information, but also to form solid tumors of primary cancer cells for toxicology studies. / <p>QC 20150113</p>

No description available.