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1.
Curr Biol ; 2021 Sep 03.
Artigo em Inglês | MEDLINE | ID: mdl-34506730

RESUMO

A fundamental question in neuroscience is whether neuronal circuits with variable circuit parameters that produce similar outputs respond comparably to equivalent perturbations.1-4 Work on the pyloric rhythm of the crustacean stomatogastric ganglion (STG) showed that highly variable sets of intrinsic and synaptic conductances can generate similar circuit activity patterns.5-9 Importantly, in response to physiologically relevant perturbations, these disparate circuit solutions can respond robustly and reliably,10-12 but when exposed to extreme perturbations the underlying circuit parameter differences produce diverse patterns of disrupted activity.7,12,13 In this example, the pyloric circuit is unchanged; only the conductance values vary. In contrast, the gastric mill rhythm in the STG can be generated by distinct circuits when activated by different modulatory neurons and/or neuropeptides.14-21 Generally, these distinct circuits produce different gastric mill rhythms. However, the rhythms driven by stimulating modulatory commissural neuron 1 (MCN1) and bath-applying CabPK (Cancer borealis pyrokinin) peptide generate comparable output patterns, despite having distinct circuits that use separate cellular and synaptic mechanisms.22-25 Here, we use these two gastric mill circuits to determine whether such circuits respond comparably when challenged with persisting (hormonal: CCAP) or acute (sensory: GPR neuron) metabotropic influences. Surprisingly, the hormone-mediated action separates these two rhythms despite activating the same ionic current in the same circuit neuron during both rhythms, whereas the sensory neuron evokes comparable responses despite acting via different synapses during each rhythm. These results highlight the need for caution when inferring the circuit response to a perturbation when that circuit is not well defined physiologically.

2.
Eur J Immunol ; 2021 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-34505280

RESUMO

The detection of tumor-specific T cells in solid tumors is integral to interrogate endogenous antitumor responses and to advance downstream therapeutic applications. Multiple biomarkers are reported to identify endogenous tumor-specific tumor-infiltrating lymphocytes (TILs), namely CD137, PD-1, CD103, and CD39; however, a direct comparison of these molecules has yet to be performed. We evaluated these biomarkers in primary human ovarian tumor samples using single-cell mass cytometry to compare their relative phenotypic profiles, and examined their response to autologous tumor cells ex vivo. PD-1+ , CD103+ , and CD39+ TILs all contain a CD137+ cell subset, while CD137+ TILs highly co-express the aforementioned markers. CD137+ TILs exhibit the highest expression of cytotoxic effector molecules compared to PD-1+ , CD103+ , or CD39+ TILs. Removal of CD137+ cells from PD-1+ , CD103+ , or CD39+ TILs diminish their IFN-γ secretion in response to autologous tumor cell stimulation, while CD137+ TILs maintain high HLA-dependent IFN-γ secretion. CD137+ TILs exhibited an exhausted phenotype but with CD28 co-expression, suggesting possible receptiveness to reinvigoration via immune checkpoint blockade. Together, our findings demonstrate that the antitumor abilities of PD-1+ , CD103+ , and CD39+ TILs are mainly derived from a subset of CD137-expressing TILs, implicating CD137 as a more selective biomarker for naturally occurring tumor-specific TILs.

3.
Blood Cancer J ; 11(8): 146, 2021 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-34392311

RESUMO

CAR-T-cell therapy against MM currently shows promising results, but usually with serious toxicities. CAR-NK cells may exert less toxicity when redirected against resistant myeloma cells. CARs can be designed through the use of receptors, such as NKG2D, which recognizes a wide range of ligands to provide broad target specificity. Here, we test this approach by analyzing the antitumor activity of activated and expanded NK cells (NKAE) and CD45RA- T cells from MM patients that were engineered to express an NKG2D-based CAR. NKAE cells were cultured with irradiated Clone9.mbIL21 cells. Then, cells were transduced with an NKG2D-4-1BB-CD3z-CAR. CAR-NKAE cells exhibited no evidence of genetic abnormalities. Although memory T cells were more stably transduced, CAR-NKAE cells exhibited greater in vitro cytotoxicity against MM cells, while showing minimal activity against healthy cells. In vivo, CAR-NKAE cells mediated highly efficient abrogation of MM growth, and 25% of the treated mice remained disease free. Overall, these results demonstrate that it is feasible to modify autologous NKAE cells from MM patients to safely express a NKG2D-CAR. Additionally, autologous CAR-NKAE cells display enhanced antimyeloma activity demonstrating that they could be an effective strategy against MM supporting the development of NKG2D-CAR-NK-cell therapy for MM.

4.
Med Teach ; : 1-8, 2021 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-34224286

RESUMO

PURPOSE: Ward rounds play a crucial role in the delivery of patient care in inpatient settings, but involve a complex mix of tasks, skills and challenges for junior doctors to negotiate. This study informs the development of high-quality training by identifying the activities that junior doctors perform, and those associated with stress during real-life ward rounds. MATERIALS AND METHODS: All activities performed by FY1 doctors (n = 60) over 2 ward rounds were coded in real-time by a trained observer using the work observation method by activity timing (WOMBAT). Doctors' heart rate was continuously recorded and non-metabolic peaks in heart rate used as a physiological indicator of stress. RESULTS: During ward rounds, FY1 doctors commonly engaged in indirect patient care, professional communication, documentation and observation. Very little time was spent on direct patient care (6%) or explicit supervision/education (0.01%). Heart rate data indicated that stress was highest during administrative tasks while interacting directly with patients while stepping out of rounds to complete personal tasks, when answering bleeps and while multi-tasking. CONCLUSIONS: Training that specifically covers the activities involved, skills required, and challenges inherent in real-life ward rounds may better prepare FY doctors for this complex area of practice.

5.
Nat Commun ; 12(1): 4365, 2021 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-34272369

RESUMO

Activating RAS missense mutations are among the most prevalent genomic alterations observed in human cancers and drive oncogenesis in the three most lethal tumor types. Emerging evidence suggests mutant KRAS (mKRAS) may be targeted immunologically, but mKRAS epitopes remain poorly defined. Here we employ a multi-omics approach to characterize HLA class I-restricted mKRAS epitopes. We provide proteomic evidence of mKRAS epitope processing and presentation by high prevalence HLA class I alleles. Select epitopes are immunogenic enabling mKRAS-specific TCRαß isolation. TCR transfer to primary CD8+ T cells confers cytotoxicity against mKRAS tumor cell lines independent of histologic origin, and the kinetics of lytic activity correlates with mKRAS peptide-HLA class I complex abundance. Adoptive transfer of mKRAS-TCR engineered CD8+ T cells leads to tumor eradication in a xenograft model of metastatic lung cancer. This study validates mKRAS peptides as bona fide epitopes facilitating the development of immune therapies targeting this oncoprotein.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Carcinogênese/imunologia , Epitopos de Linfócito T/imunologia , Neoplasias Pulmonares/imunologia , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Transferência Adotiva , Alelos , Animais , Carcinogênese/genética , Linhagem Celular Tumoral , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Camundongos , Mutação , Peptídeos/genética , Peptídeos/imunologia , Proteômica , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Ensaios Antitumorais Modelo de Xenoenxerto
6.
J Immunother Cancer ; 9(7)2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34253637

RESUMO

BACKGROUND: Bispecific T cell engagers represent the majority of bispecific antibodies (BsAbs) entering the clinic to treat metastatic cancer. The ability to apply these agents safely and efficaciously in the clinic, particularly for solid tumors, has been challenging. Many preclinical studies have evaluated parameters related to the activity of T cell engaging BsAbs, but many questions remain. MAIN BODY: This study investigates the impact of affinity of T cell engaging BsAbs with regards to potency, efficacy, and induction of immunomodulatory receptors/ligands using HER-2/CD3 BsAbs as a model system. We show that an IgG BsAb can be as efficacious as a smaller BsAb format both in vitro and in vivo. We uncover a dichotomous relationship between tumor-associated antigen (TAA) affinity and CD3 affinity requirements for cells that express high versus low levels of TAA. HER-2 affinity directly correlated with the CD3 engager lysis potency of HER-2/CD3 BsAbs when HER-2 receptor numbers are high (~200 K/cell), while the CD3 affinity did not impact potency until its binding affinity was extremely low (<600 nM). When HER-2 receptor numbers were lower (~20 K/cell), both HER-2 and CD3 affinity impacted potency. The high affinity anti-HER-2/low CD3 affinity BsAb also demonstrated lower cytokine induction levels in vivo and a dosing paradigm atypical of extremely high potency T cell engaging BsAbs reaching peak efficacy at doses >3 mg/kg. This data confirms that low CD3 affinity provides an opportunity for improved safety and dosing for T cell engaging BsAbs. T cell redirection also led to upregulation of Programmed cell death 1 (PD-1) and 4-1BB, but not CTLA-4 on T cells, and to Programmed death-ligand 1 (PD-L1) upregulation on HER-2HI SKOV3 tumor cells, but not on HER-2LO OVCAR3 tumor cells. Using this information, we combined anti-PD-1 or anti-4-1BB monoclonal antibodies with the HER-2/CD3 BsAb in vivo and demonstrated significantly increased efficacy against HER-2HI SKOV3 tumors via both combinations. CONCLUSIONS: Overall, these studies provide an informational dive into the optimization process of CD3 engaging BsAbs for solid tumors indicating that a reduced affinity for CD3 may enable a better therapeutic index with a greater selectivity for the target tumor and a reduced cytokine release syndrome. These studies also provide an additional argument for combining T cell checkpoint inhibition and co-stimulation to achieve optimal efficacy.

7.
Int J Mol Sci ; 22(11)2021 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-34070369

RESUMO

Folate receptor beta (FRß) is a folate binding receptor expressed on myeloid lineage hematopoietic cells. FRß is commonly expressed at high levels on malignant blasts in patients with acute myeloid leukemia (AML), as well as on M2 polarized tumor-associated macrophages (TAMs) in the tumor microenvironment of many solid tumors. Therefore, FRß is a potential target for both direct and indirect cancer therapy. We demonstrate that FRß is expressed in both AML cell lines and patient-derived AML samples and that a high-affinity monoclonal antibody against FRß (m909) has the ability to cause dose- and expression-dependent ADCC against these cells in vitro. Importantly, we find that administration of m909 has a significant impact on tumor growth in a humanized mouse model of AML. Surprisingly, m909 functions in vivo with and without the infusion of human NK cells as mediators of ADCC, suggesting potential involvement of mouse macrophages as effector cells. We also found that TAMs from primary ovarian ascites samples expressed appreciable levels of FRß and that m909 has the ability to cause ADCC in these samples. These results indicate that the targeting of FRß using m909 has the potential to limit the outgrowth of AML in vitro and in vivo. Additionally, m909 causes cytotoxicity to TAMs in the tumor microenvironment of ovarian cancer warranting further investigation of m909 and its derivatives as therapeutic agents in patients with FRß-expressing cancers.


Assuntos
Antineoplásicos Imunológicos/farmacologia , Receptor 2 de Folato , Imunoterapia , Leucemia Mieloide Aguda , Proteínas de Neoplasias , Neoplasias Ovarianas , Animais , Células CHO , Cricetulus , Feminino , Receptor 2 de Folato/antagonistas & inibidores , Receptor 2 de Folato/imunologia , Células HL-60 , Humanos , Leucemia Mieloide Aguda/imunologia , Leucemia Mieloide Aguda/patologia , Leucemia Mieloide Aguda/terapia , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/imunologia , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/patologia , Neoplasias Ovarianas/terapia , Células THP-1 , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Nat Commun ; 12(1): 877, 2021 02 09.
Artigo em Inglês | MEDLINE | ID: mdl-33563975

RESUMO

The immunosuppressive tumor microenvironment (TME) represents a major barrier for effective immunotherapy. Tumor-associated macrophages (TAMs) are highly heterogeneous and plastic cell components of the TME which can either promote tumor progression (M2-like) or boost antitumor immunity (M1-like). Here, we demonstrate that a subset of TAMs that express folate receptor ß (FRß) possess an immunosuppressive M2-like profile. In syngeneic tumor mouse models, chimeric antigen receptor (CAR)-T cell-mediated selective elimination of FRß+ TAMs in the TME results in an enrichment of pro-inflammatory monocytes, an influx of endogenous tumor-specific CD8+ T cells, delayed tumor progression, and prolonged survival. Preconditioning of the TME with FRß-specific CAR-T cells also improves the effectiveness of tumor-directed anti-mesothelin CAR-T cells, while simultaneous co-administration of both CAR products does not. These results highlight the pro-tumor role of FRß+ TAMs in the TME and the therapeutic implications of TAM-depleting agents as preparative adjuncts to conventional immunotherapies that directly target tumor antigens.


Assuntos
Imunoterapia Adotiva/métodos , Neoplasias/terapia , Receptores de Antígenos Quiméricos/imunologia , Macrófagos Associados a Tumor/imunologia , Animais , Linfócitos T CD8-Positivos/imunologia , Receptor 2 de Folato/imunologia , Receptor 2 de Folato/metabolismo , Humanos , Imunossupressão , Camundongos , Camundongos Endogâmicos C57BL , Monócitos/imunologia , Neoplasias/imunologia , Células Tumorais Cultivadas , Microambiente Tumoral/imunologia , Macrófagos Associados a Tumor/metabolismo
9.
Molecules ; 25(24)2020 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-33352773

RESUMO

Theranostics are emerging as a pillar of cancer therapy that enable the use of single molecule constructs for diagnostic and therapeutic application. As poly adenosine diphosphate (ADP)-ribose polymerase 1 (PARP-1) is overexpressed in various cancer types, and is localized to the nucleus, PARP-1 can be safely targeted with Auger emitters to induce DNA damage in tumors. Here, we investigated a radioiodinated PARP inhibitor, [125I]KX1, and show drug target specific DNA damage and subsequent killing of BRCA1 and non-BRCA mutant ovarian cancer cells at sub-pharmacological concentrations several orders of magnitude lower than traditional PARP inhibitors. Furthermore, we demonstrated that viable tumor tissue from ovarian cancer patients can be used to screen tumor radiosensitivity ex-vivo, enabling the direct assessment of therapeutic efficacy. Finally, we showed tumors can be imaged by single-photon computed tomography (SPECT) with PARP theranostic, [123I]KX1, in a human ovarian cancer xenograft mouse model. These data support the utility of PARP-1 targeted radiopharmaceutical therapy as a theranostic option for PARP-1 overexpressing ovarian cancers.


Assuntos
Antineoplásicos/farmacologia , Proteína BRCA1/genética , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/genética , Poli(ADP-Ribose) Polimerase-1/genética , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Animais , Linhagem Celular Tumoral , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/genética , Modelos Animais de Doenças , Feminino , Xenoenxertos , Humanos , Radioisótopos do Iodo/farmacologia , Camundongos SCID
10.
Artigo em Inglês | MEDLINE | ID: mdl-32923884

RESUMO

PURPOSE: Homologous DNA repair-deficient (HRD) ovarian cancers (OCs), including those with BRCA1/2 mutations, have higher levels of genetic instability, potentially resulting in higher immunogenicity, and have been suggested to respond better to immune checkpoint inhibitors (ICIs) than homologous DNA repair-proficient OCs. However, clinical evidence is lacking. The study aimed to evaluate the associations between BRCA1/2 mutations, HRD, and other genomic parameters and response to ICIs and survival in OC. METHODS: This is a single-institution retrospective analysis of women with recurrent OC treated with ICIs. BRCA1/2 mutation status and clinicopathologic variables were abstracted from the medical records. Targeted and whole-exome sequencing data available for a subset of patients were used to assess tumor mutational burden (TMB), HRD, and fraction of genome altered (FGA). ICI response was defined as lack of disease progression for ≥ 24 weeks. Associations of BRCA1/2 status and genomic alterations with progression-free survival (PFS) and overall survival (OS) were determined using Cox proportional hazards models. RESULTS: Of the 143 women treated with ICIs, 134 had known BRCA1/2 mutation status. Deleterious germline or somatic BRCA1/2 mutations were present in 31 women (24%). There was no association between presence of BRCA1/2 mutations and response (P = .796) or survival. Genomic analysis in 73 women found no association between TMB (P = .344) or HRD (P = .222) and response, PFS, or OS. There were also no significant differences in somatic genetic alterations between responders and nonresponders. High FGA was associated with an improvement in PFS (P = .014) and OS (P = .01). CONCLUSION: TMB, BRCA1/2 mutations, and HRD are not associated with response or survival, cautioning against their use as selection criteria for ICI in recurrent OC. FGA should be investigated further as a biomarker of response to immunotherapy in OC.

11.
Front Immunol ; 11: 1109, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32625204

RESUMO

Chimeric antigen receptor (CAR) T cell therapies have demonstrated remarkable efficacy for the treatment of hematological malignancies. However, in patients with solid tumors, objective responses to CAR-T cell therapy remain sporadic and transient. A major obstacle for CAR-T cells is the intrinsic ability of tumors to evade immune responses. Advanced solid tumors are largely composed of desmoplastic stroma and immunosuppressive modulators, and characterized by aberrant cell proliferation and vascularization, resulting in hypoxia and altered nutrient availability. To mount a curative response after infusion, CAR-T cells must infiltrate the tumor, recognize their cognate antigen and perform their effector function in this hostile tumor microenvironment, to then differentiate and persist as memory T cells that confer long-term protection. Fortunately, recent advances in synthetic biology provide a wide set of tools to genetically modify CAR-T cells to overcome some of these obstacles. In this review, we provide a comprehensive overview of the key tumor intrinsic mechanisms that prevent an effective CAR-T cell antitumor response and we discuss the most promising strategies to prevent tumor escape to CAR-T cell therapy.


Assuntos
Imunoterapia Adotiva , Neoplasias/imunologia , Neoplasias/terapia , Receptores de Antígenos Quiméricos/imunologia , Evasão Tumoral/imunologia , Microambiente Tumoral/imunologia , Animais , Matriz Extracelular/imunologia , Fibroblastos/imunologia , Humanos , Proteínas de Checkpoint Imunológico/imunologia , Linfócitos do Interstício Tumoral/imunologia , Macrófagos/imunologia , Camundongos , Modelos Imunológicos , Células Supressoras Mieloides/imunologia , Neoplasias/irrigação sanguínea , Linfócitos T Reguladores/imunologia , Hipóxia Tumoral/imunologia
12.
Int J Behav Nutr Phys Act ; 17(1): 86, 2020 07 06.
Artigo em Inglês | MEDLINE | ID: mdl-32631366

RESUMO

BACKGROUND: Hospitals in the UK offer snacks for sale to patients, staff and visitors. Despite the NHS's health promoting role, and tightening of regulations around which foods can be sold in hospitals, many snacks purchased in this setting are unhealthy. The present project tests the effectiveness of theory-based point of purchase prompts (PPPs; a form of cognitive nudge) designed to make it cognitively easier for consumers to compare available products and choose healthier options. METHODS: Hospital shops in Scotland (n = 30) were recruited into a cluster randomised controlled trial to test whether a PPP could reduce the average calorie, fat and/or sugar content of purchased snacks. Inclusion criteria stated that eligible sites; sold food; were located in a hospital; and were accessible to staff, patients and visitors. The PPP intervention was a theory-based sign (tailored to the available range in each location) designed to cognitively simplify healthier snack choices by facilitating cross-product comparison. Shops were randomised to display PPPs (intervention; n = 15) or not (control; n = 15) using block randomisation controlling for shop size. Data on all snacks purchased from participating shops were obtained from retailers for a 12-week baseline and 12-week follow-up period. Primary outcomes were the average calorie (kcals), fat(g) and sugar(g) content of snacks purchased each day. Secondary outcomes were the average customer spend per item purchased (£,p) and the total number of snacks purchased daily. Shop staff were not blinded to group assignment but data providers were. Data were analysed using mixed effects multi-level regression models. RESULTS: Data from > 1 million snack purchases were analysed. Snacks purchased from intervention sites were on average significantly lower in calorie (γ = - 1.84, p < .001) and sugar (γ = - 0.18, p = .030) at follow up relative to baseline but only the reduction in calories was significantly different to control. Average spend per item also reduced significantly in intervention (but not control) sites (γ = - 0.89, p < .001). The intervention had no effect on the fat content of snacks or the number of snacks sold. CONCLUSIONS: Simple, theory-based point of purchase prompts can produce small but statistically significant reductions in the energy content of snack purchases from hospital shops. TRIAL REGISTRATION: Retrospectively registered (8/Oct/2018) with ISRCTN (ID: ISRCTN90365793 ).


Assuntos
Comércio/estatística & dados numéricos , Comportamento do Consumidor/estatística & dados numéricos , Dieta Saudável , Preferências Alimentares/psicologia , Lojas no Hospital , Lanches/psicologia , Gorduras na Dieta , Açúcares da Dieta , Ingestão de Energia , Promoção da Saúde , Humanos , Valor Nutritivo , Escócia
13.
J Clin Oncol ; 38(16): 1814-1823, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32275468

RESUMO

PURPOSE: Single-agent PD-1 blockade exhibits limited efficacy in epithelial ovarian cancer (EOC). We evaluated ipilimumab plus nivolumab compared with nivolumab alone in women with persistent or recurrent EOC. METHODS: Eligibility criteria included measurable disease, 1-3 prior regimens, and platinum-free interval (PFI) < 12 months. Participants were randomly allocated to intravenous nivolumab (every 2 weeks) or induction with nivolumab plus ipilimumab for 4 doses (every 3 weeks), followed by every-2-week maintenance nivolumab for a maximum of 42 doses. The primary null hypothesis was equal probability of objective response within 6 months of random allocation in each arm. RESULTS: One hundred patients were allocated to receive either nivolumab (n = 49), or nivolumab plus ipilimumab (n = 51), with PFI of < 6 months in 62%. Six (12.2%) responses occurred within 6 months in the nivolumab group and 16 (31.4%) in the nivolumab plus ipilimumab group (odds ratio, 3.28; 85% CI, 1.54 to infinity; P = .034). The median progression-free survival (PFS) was 2 and 3.9 months in the nivolumab and nivolumab plus ipilimumab groups, respectively, with a PFI-stratified hazard ratio of 0.53 (95% CI, 0.34 to 0.82); the respective hazard ratio for death was 0.79 (95% CI, 0.44 to 1.42). Grade ≥ 3 related adverse events occurred in 33% of patients in the nivolumab group and 49% in the combination group, with no treatment-related deaths. PD-L1 expression was not significantly associated with response in either treatment group. CONCLUSION: Compared with nivolumab alone, the combination of nivolumab and ipilimumab in EOC resulted in superior response rate and longer, albeit limited, PFS, with toxicity of the combination regimen comparable to prior reports. Additional combination studies to enhance durability of the dual regimen are warranted.


Assuntos
Antineoplásicos Imunológicos/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma Epitelial do Ovário/tratamento farmacológico , Inibidores de Checkpoint Imunológico/administração & dosagem , Ipilimumab/administração & dosagem , Recidiva Local de Neoplasia , Nivolumabe/administração & dosagem , Neoplasias Ovarianas/tratamento farmacológico , Adulto , Idoso , Idoso de 80 Anos ou mais , Antineoplásicos Imunológicos/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Carcinoma Epitelial do Ovário/imunologia , Carcinoma Epitelial do Ovário/mortalidade , Feminino , Humanos , Inibidores de Checkpoint Imunológico/efeitos adversos , Ipilimumab/efeitos adversos , Pessoa de Meia-Idade , Nivolumabe/efeitos adversos , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/mortalidade , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Intervalo Livre de Progressão , Fatores de Tempo , Estados Unidos
14.
J Neurophysiol ; 123(5): 2075-2089, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32319837

RESUMO

Elevated potassium concentration ([K+]) is often used to alter excitability in neurons and networks by shifting the potassium equilibrium potential (EK) and, consequently, the resting membrane potential. We studied the effects of increased extracellular [K+] on the well-described pyloric circuit of the crab Cancer borealis. A 2.5-fold increase in extracellular [K+] (2.5×[K+]) depolarized pyloric dilator (PD) neurons and resulted in short-term loss of their normal bursting activity. This period of silence was followed within 5-10 min by the recovery of spiking and/or bursting activity during continued superfusion of 2.5×[K+] saline. In contrast, when PD neurons were pharmacologically isolated from pyloric presynaptic inputs, they exhibited no transient loss of spiking activity in 2.5×[K+], suggesting the presence of an acute inhibitory effect mediated by circuit interactions. Action potential threshold in PD neurons hyperpolarized during an hour-long exposure to 2.5×[K+] concurrent with the recovery of spiking and/or bursting activity. Thus the initial loss of activity appears to be mediated by synaptic interactions within the network, but the secondary adaptation depends on changes in the intrinsic excitability of the pacemaker neurons. The complex sequence of events in the responses of pyloric neurons to elevated [K+] demonstrates that electrophysiological recordings are necessary to determine both the transient and longer term effects of even modest alterations of K+ concentrations on neuronal activity.NEW & NOTEWORTHY Solutions with elevated extracellular potassium are commonly used as a depolarizing stimulus. We studied the effects of high potassium concentration ([K+]) on the pyloric circuit of the crab stomatogastric ganglion. A 2.5-fold increase in extracellular [K+] caused a transient loss of activity that was not due to depolarization block, followed by a rapid increase in excitability and recovery of spiking within minutes. This suggests that changing extracellular potassium can have complex and nonstationary effects on neuronal circuits.


Assuntos
Braquiúros/fisiologia , Geradores de Padrão Central/fisiologia , Fenômenos Eletrofisiológicos/fisiologia , Gânglios dos Invertebrados/fisiologia , Potássio/metabolismo , Piloro/fisiologia , Animais , Geradores de Padrão Central/metabolismo , Gânglios dos Invertebrados/metabolismo , Masculino , Piloro/metabolismo
15.
J Am Chem Soc ; 142(14): 6554-6568, 2020 04 08.
Artigo em Inglês | MEDLINE | ID: mdl-32191035

RESUMO

Universal immune receptors represent a rapidly emerging form of adoptive T-cell therapy with the potential to overcome safety and antigen escape challenges faced by conventional chimeric antigen receptor (CAR) T-cell therapy. By decoupling antigen recognition and T-cell signaling domains via bifunctional antigen-specific targeting ligands, universal immune receptors can regulate T-cell effector function and target multiple antigens with a single receptor. Here, we describe the development of the SpyCatcher immune receptor, the first universal immune receptor that allows for the post-translational covalent attachment of targeting ligands at the T-cell surface through the application of SpyCatcher-SpyTag chemistry. The SpyCatcher immune receptor redirected primary human T cells against a variety of tumor antigens via the addition of SpyTag-labeled targeting ligands, both in vitro and in vivo. SpyCatcher T-cell activity relied upon the presence of both target antigen and SpyTag-labeled targeting ligand, allowing for dose-dependent control of function. The mutational disruption of covalent bond formation between the receptor and the targeting ligand still permitted redirected T-cell function but significantly compromised antitumor function. Thus, the SpyCatcher immune receptor allows for rapid antigen-specific receptor assembly, multiantigen targeting, and controllable T-cell activity.


Assuntos
Engenharia Genética/métodos , Linfócitos T/imunologia , Humanos , Ligantes
16.
Gynecol Oncol ; 156(1): 222-232, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31818495

RESUMO

OBJECTIVE: The aim of this study was to "humanize" ovarian cancer patient-derived xenograft (PDX) models by autologous transfer of patient-matched tumor infiltrating lymphocytes (TILs) to evaluate immunotherapies. METHODS: Orthotopic high-grade serous ovarian cancer (HGSOC) PDX models were established from three patient donors. Models were molecularly and histologically validated by immunohistochemistry. TILs were expanded from donor tumors using a rapid expansion protocol. Ex vivo TIL and tumor co-cultures were performed to validate TIL reactivity against patient-matched autologous tumor cells. Expression of TIL activation markers and cytokine secretion was quantitated by flow cytometry and ELISA. As proof of concept, the efficacy of anti-PD-1 monotherapy was tested in autologous TIL/tumor HGSOC PDX models. RESULTS: Evaluation of T-cell activation in autologous TIL/tumor co-cultures resulted in an increase in HLA-dependent IFNγ production and T-cell activation. In response to increased IFNγ production, tumor cell expression of PD-L1 was increased. Addition of anti-PD-1 antibody to TIL/tumor co-cultures increased autologous tumor lysis in a CCNE1 amplified model. Orthotopic HGSOC PDX models from parallel patient-matched tumors maintained their original morphology and molecular marker profile. Autologous tumor-reactive TIL administration in patient-matched PDX models resulted in reduced tumor burden and increased survival, in groups that also received anti-PD-1 therapy. CONCLUSIONS: This study validates a novel, clinically relevant model system for in vivo testing of immunomodulating therapeutic strategies for ovarian cancer, and provides a unique platform for assessing patient-specific T-cell response to immunotherapy.


Assuntos
Imunoterapia/métodos , Linfócitos do Interstício Tumoral/imunologia , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/terapia , Ensaios Antitumorais Modelo de Xenoenxerto/métodos , Animais , Feminino , Humanos , Ativação Linfocitária , Linfócitos do Interstício Tumoral/transplante , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Transplante de Neoplasias/métodos , Neoplasias Ovarianas/patologia , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/imunologia , Linfócitos T/imunologia
17.
Mol Ther ; 28(2): 548-560, 2020 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-31870622

RESUMO

The prognosis of patients diagnosed with advanced ovarian or endometrial cancer remains poor, and effective therapeutic strategies are limited. The Müllerian inhibiting substance type 2 receptor (MISIIR) is a transforming growth factor ß (TGF-ß) receptor family member, overexpressed by most ovarian and endometrial cancers while absent in most normal tissues. Restricted tissue expression, coupled with an understanding that MISIIR ligation transmits apoptotic signals to cancer cells, makes MISIIR an attractive target for tumor-directed therapeutics. However, the development of clinical MISIIR-targeted agents has been challenging. Prompted by the responses achieved in patients with blood malignancies using chimeric antigen receptor (CAR) T cell therapy, we hypothesized that MISIIR targeting may be achieved using a CAR T cell approach. Herein, we describe the development and evaluation of a CAR that targets MISIIR. T cells expressing the MISIIR-specific CAR demonstrated antigen-specific reactivity in vitro and eliminated MISIIR-overexpressing tumors in vivo. MISIIR CAR T cells also recognized a panel of human ovarian and endometrial cancer cell lines, and they lysed a battery of patient-derived tumor specimens in vitro, without mediating cytotoxicity of a panel of normal primary human cells. In conclusion, these results indicate that MISIIR targeting for the treatment of ovarian cancer and other gynecologic malignancies is achievable using CAR technology.


Assuntos
Neoplasias dos Genitais Femininos/imunologia , Imunoterapia Adotiva , Neoplasias Ovarianas/imunologia , Receptores de Antígenos Quiméricos/imunologia , Receptores de Peptídeos/imunologia , Receptores de Fatores de Crescimento Transformadores beta/imunologia , Linfócitos T/imunologia , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Epitopos/genética , Epitopos/imunologia , Feminino , Neoplasias dos Genitais Femininos/terapia , Humanos , Camundongos , Neoplasias Ovarianas/terapia , Receptores de Antígenos Quiméricos/metabolismo , Anticorpos de Cadeia Única/genética , Anticorpos de Cadeia Única/imunologia , Linfócitos T/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Cancer Cell ; 35(6): 885-900.e10, 2019 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-31185212

RESUMO

We investigated the role of chemokines in regulating T cell accumulation in solid tumors. CCL5 and CXCL9 overexpression was associated with CD8+ T cell infiltration in solid tumors. T cell infiltration required tumor cell-derived CCL5 and was amplified by IFN-γ-inducible, myeloid cell-secreted CXCL9. CCL5 and CXCL9 coexpression revealed immunoreactive tumors with prolonged survival and response to checkpoint blockade. Loss of CCL5 expression in human tumors was associated with epigenetic silencing through DNA methylation. Reduction of CCL5 expression caused tumor-infiltrating lymphocyte (TIL) desertification, whereas forced CCL5 expression prevented Cxcl9 expression and TILs loss, and attenuated tumor growth in mice through IFN-γ. The cooperation between tumor-derived CCL5 and IFN-γ-inducible CXCR3 ligands secreted by myeloid cells is key for orchestrating T cell infiltration in immunoreactive and immunoresponsive tumors.


Assuntos
Linfócitos T CD8-Positivos/metabolismo , Quimiotaxia de Leucócito , Citocinas/metabolismo , Células Dendríticas/metabolismo , Ativação Linfocitária , Linfócitos do Interstício Tumoral/metabolismo , Macrófagos/metabolismo , Neoplasias Ovarianas/metabolismo , Animais , Antineoplásicos Imunológicos/farmacologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular Tumoral , Quimiocina CCL5/genética , Quimiocina CCL5/imunologia , Quimiocina CCL5/metabolismo , Quimiocina CXCL9/genética , Quimiocina CXCL9/imunologia , Quimiocina CXCL9/metabolismo , Quimiotaxia de Leucócito/efeitos dos fármacos , Técnicas de Cocultura , Citocinas/genética , Citocinas/imunologia , Metilação de DNA , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Epigênese Genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Imunoterapia/métodos , Interferon gama/genética , Interferon gama/imunologia , Interferon gama/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/imunologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos Endogâmicos C57BL , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/patologia , Neoplasias Ovarianas/terapia , Comunicação Parácrina , Receptores CXCR3/genética , Receptores CXCR3/imunologia , Receptores CXCR3/metabolismo , Transdução de Sinais
19.
Front Oncol ; 9: 176, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30984613

RESUMO

Chimeric antigen receptor (CAR) T cells have shown great success in the treatment of CD19+ hematological malignancies, leading to their recent approval by the FDA as a new cancer treatment modality. However, their broad use is limited since a CAR targets a single tumor associated antigen (TAA), which is not effective against tumors with heterogeneous TAA expression or emerging antigen loss variants. Further, stably engineered CAR T cells can continually and uncontrollably proliferate and activate in response to antigen, potentially causing fatal on-target off-tumor toxicity, cytokine release syndrome, or neurotoxicity without a method of control or elimination. To address these issues, our lab and others have developed various universal immune receptors (UIRs) that allow for targeting of multiple TAAs by T cells expressing a single receptor. UIRs function through the binding of an extracellular adapter domain which acts as a bridge between intracellular T cell signaling domains and a soluble tumor antigen targeting ligand (TL). The dissociation of TAA targeting and T cell signaling confers many advantages over standard CAR therapy, such as dose control of T cell effector function, the ability to simultaneously or sequentially target multiple TAAs, and control of immunologic synapse geometry. There are currently four unique UIR platform types: ADCC-mediating Fc-binding immune receptors, bispecific protein engaging immune receptors, natural binding partner immune receptors, and anti-tag CARs. These UIRs all allow for potential benefits over standard CARs, but also bring unique engineering challenges that will have to be addressed to achieve maximal efficacy and safety in the clinic. Still, UIRs present an exciting new avenue for adoptive T cell transfer therapies and could lead to their expanded use in areas which current CAR therapies have failed. Here we review the development of each UIR platform and their unique functional benefits, and detail the potential hurdles that may need to be overcome for continued clinical translation.

20.
Cancer ; 125(12): 1963-1972, 2019 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-30835824

RESUMO

Substantial progress has been made in understanding ovarian cancer at the molecular and cellular level. Significant improvement in 5-year survival has been achieved through cytoreductive surgery, combination platinum-based chemotherapy, and more effective treatment of recurrent cancer, and there are now more than 280,000 ovarian cancer survivors in the United States. Despite these advances, long-term survival in late-stage disease has improved little over the last 4 decades. Poor outcomes relate, in part, to late stage at initial diagnosis, intrinsic drug resistance, and the persistence of dormant drug-resistant cancer cells after primary surgery and chemotherapy. Our ability to accelerate progress in the clinic will depend on the ability to answer several critical questions regarding this disease. To assess current answers, an American Association for Cancer Research Special Conference on "Critical Questions in Ovarian Cancer Research and Treatment" was held in Pittsburgh, Pennsylvania, on October 1-3, 2017. Although clinical, translational, and basic investigators conducted much of the discussion, advocates participated in the meeting, and many presentations were directly relevant to patient care, including treatment with poly adenosine diphosphate ribose polymerase (PARP) inhibitors, attempts to improve immunotherapy by overcoming the immune suppressive effects of the microenvironment, and a better understanding of the heterogeneity of the disease.


Assuntos
Antineoplásicos/uso terapêutico , Imunoterapia/métodos , Neoplasias Ovarianas/diagnóstico , Neoplasias Ovarianas/tratamento farmacológico , Assistência Centrada no Paciente , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Congressos como Assunto , Resistencia a Medicamentos Antineoplásicos , Feminino , Humanos , Sociedades Científicas , Microambiente Tumoral
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