Personalizing non-small cell lung cancer treatment through patient-derived xenograft models: preclinical and clinical factors for consideration.

PURPOSE: In the pursuit of creating personalized and more effective treatment strategies for lung cancer patients, Patient-Derived Xenografts (PDXs) have been introduced as preclinical platforms that can recapitulate the specific patient's tumor in an in vivo model. We investigated how well PDX models can preserve the tumor's clinical and molecular characteristics across different generations. METHODS: A Non-Small Cell Lung Cancer (NSCLC) PDX model was established in NSG-SGM3 mice and clinical and preclinical factors were assessed throughout subsequent passages. Our cohort consisted of 40 NSCLC patients, which were used to create 20 patient-specific PDX models in NSG-SGM3 mice. Histopathological staining and Whole Exome Sequencing (WES) analysis were preformed to understand tumor heterogeneity throughout serial passages. RESULTS: The main factors that contributed to the growth of the engrafted PDX in mice were a higher grade or stage of disease, in contrast to the long duration of chemotherapy treatment, which was negatively correlated with PDX propagation. Successful PDX growth was also linked to poorer prognosis and overall survival, while growth pattern variability was affected by the tumor aggressiveness, primarily affecting the first passage. Pathology analysis showed preservation of the histological type and grade; however, WES analysis revealed genomic instability in advanced passages, leading to the inconsistencies in clinically relevant alterations between the PDXs and biopsies. CONCLUSIONS: Our study highlights the impact of multiple clinical and preclinical factors on the engraftment success, growth kinetics, and tumor stability of patient-specific NSCLC PDXs, and underscores the importance of considering these factors when guiding and evaluating prolonged personalized treatment studies for NSCLC patients in these models, as well as signaling the imperative for additional investigations to determine the full clinical potential of this technique.

Knockdown of Antisense Noncoding Mitochondrial RNA Reduces Tumorigenicity of Patient-Derived Clear Cell Renal Carcinoma Cells in an Orthotopic Xenograft Mouse Model.

Clear cell renal cell carcinoma (ccRCC) is the most prevalent form of renal cancer and its treatment is hindered by a resistance to targeted therapies, immunotherapies and combinations of both. We have reported that the knockdown of the antisense noncoding mitochondrial RNAs (ASncmtRNAs) with chemically modified antisense oligonucleotides induces proliferative arrest and apoptotic death in tumor cells from many human and mouse cancer types. These studies have been mostly performed in vitro and in vivo on commercially available cancer cell lines and have shown that in mouse models tumor growth is stunted by the treatment. The present work was performed on cells derived from primary and metastatic ccRCC tumors. We established primary cultures from primary and metastatic ccRCC tumors, which were subjected to knockdown of ASncmtRNAs in vitro and in vivo in an orthotopic xenograft model in NOD/SCID mice. We found that these primary ccRCC cells are affected in the same way as tumor cell lines and in the orthotopic model tumor growth was significantly reduced by the treatment. This study on patient-derived ccRCC tumor cells represents a model closer to actual patient ccRCC tumors and shows that knockdown of ASncmtRNAs poses a potential treatment option for these patients.

Advances towards the use of gastrointestinal tumor patient-derived organoids as a therapeutic decision-making tool.

In December 2022 the US Food and Drug Administration (FDA) removed the requirement that drugs in development must undergo animal testing before clinical evaluation, a declaration that now demands the establishment and verification of ex vivo preclinical models that closely represent tumor complexity and that can predict therapeutic response. Fortunately, the emergence of patient-derived organoid (PDOs) culture has enabled the ex vivo mimicking of the pathophysiology of human tumors with the reassembly of tissue-specific features. These features include histopathological variability, molecular expression profiles, genetic and cellular heterogeneity of parental tissue, and furthermore growing evidence suggests the ability to predict patient therapeutic response. Concentrating on the highly lethal and heterogeneous gastrointestinal (GI) tumors, herein we present the state-of-the-art and the current methodology of PDOs. We highlight the potential additions, improvements and testing required to allow the ex vivo of study the tumor microenvironment, as well as offering commentary on the predictive value of clinical response to treatments such as chemotherapy and immunotherapy.

Patient-Derived Xenotransplant of CNS Neoplasms in Zebrafish: A Systematic Review.

Glioblastoma and neuroblastoma are the most common central nervous system malignant tumors in adult and pediatric populations. Both are associated with poor survival. These tumors are highly heterogeneous, having complex interactions among different cells within the tumor and with the tumor microenvironment. One of the main challenges in the neuro-oncology field is achieving optimal conditions to evaluate a tumor's molecular genotype and phenotype. In this respect, the zebrafish biological model is becoming an excellent alternative for studying carcinogenic processes and discovering new treatments. This review aimed to describe the results of xenotransplantation of patient-derived CNS tumors in zebrafish models. The reviewed studies show that it is possible to maintain glioblastoma and neuroblastoma primary cell cultures and transplant the cells into zebrafish embryos. The zebrafish is a suitable biological model for understanding tumor progression and the effects of different treatments. This model offers new perspectives in providing personalized care and improving outcomes for patients living with central nervous system tumors.

Central giant cell granulomas of the jaws stromal cells harbour mutations and have osteogenic differentiation capacity, in vivo and in vitro.

BACKGROUND: Central giant cell granulomas (CGCG) of the jaws are osteolytic lesions that may behave aggressively and respond poorly to surgery. Microscopically, in addition to giant cells, there is a mononuclear cell population composed of macrophage/monocytic cells and spindle-shaped cells of mesenchymal origin. Seventy two percent of these tumours harbour mutually exclusive TRPV4, KRAS and FGFR1 mutations. We aimed to assess the mutational status of mononuclear and giant cells and the osteogenic potential of stromal cells in vitro and in vivo. METHODS AND RESULTS: We screened CGCG for signature mutations and used laser-capture microdissection to demonstrate that the mutations are restricted to the mononuclear cells. Additionally, we established CGCG primary cell culture and observed that the cells retained the mutations throughout passages. By flow cytometry, we observed predominance of CD14- CD51- CD61- cells, consistent with the expected profile for stromal cells. Considering the mesenchymal origin of stromal cells, we assessed the osteogenic differentiation potential of CGCG cells in culture by cytochemistry (von Kossa and alizarin red staining), alkaline phosphatase (ALP) activity assay and gene expression of osteogenic markers. CGCG cells presented self-capacity to increase ALP levels in a time-dependent manner and under osteogenic induction presented increasing number of calcium deposits, and overall higher expression of osteocalcin, RUNX2, ALPL and osteopontin than cells without osteogenic induction. A patient-derived xenograft model for CGCG was established, and osteoid material deposition was observed. CONCLUSION: Collectively, the results confirm that the signature mutations are restricted to stromal cells in CGCG, and the in vitro and in vivo results support that these cells have the capacity to differentiate into osteoblasts, in line with the bone formation often observed in the stroma of these lesions.

In-depth characterization of a new patient-derived xenograft model for metaplastic breast carcinoma to identify viable biologic targets and patterns of matrix evolution within rare tumor types.

Metaplastic breast carcinoma (MBC) is a rare breast cancer subtype with rapid growth, high rates of metastasis, recurrence and drug resistance, and diverse molecular and histological heterogeneity. Patient-derived xenografts (PDXs) provide a translational tool and physiologically relevant system to evaluate tumor biology of rare subtypes. Here, we provide an in-depth comprehensive characterization of a new PDX model for MBC, TU-BcX-4IC. TU-BcX-4IC is a clinically aggressive tumor exhibiting rapid growth in vivo, spontaneous metastases, and elevated levels of cell-free DNA and circulating tumor cell DNA. Relative chemosensitivity of primary cells derived from TU-BcX-4IC was performed using the National Cancer Institute (NCI) oncology drug set, crystal violet staining, and cytotoxic live/dead immunofluorescence stains in adherent and organoid culture conditions. We employed novel spheroid/organoid incubation methods (Pu·MA system) to demonstrate that TU-BcX-4IC is resistant to paclitaxel. An innovative physiologically relevant system using human adipose tissue was used to evaluate presence of cancer stem cell-like populations ex vivo. Tissue decellularization, cryogenic-scanning electron microscopy imaging and rheometry revealed consistent matrix architecture and stiffness were consistent despite serial transplantation. Matrix-associated gene pathways were essentially unchanged with serial passages, as determined by qPCR and RNA sequencing, suggesting utility of decellularized PDXs for in vitro screens. We determined type V collagen to be present throughout all serial passage of TU-BcX-4IC tumor, suggesting it is required for tumor maintenance and is a potential viable target for MBC. In this study we introduce an innovative and translational model system to study cell-matrix interactions in rare cancer types using higher passage PDX tissue.

Identificação e Caracterização Funcional de Alterações Genéticas em Sarcomas Pleomórficos Indiferenciados e Fusocelulares / Identification and Functional Characterization of Genetic Alterations in Undifferentiated and Fusocellular Pleomorphic Sarcomas

Os sarcomas de partes moles (SPM) são tumores malignos derivados de células mesenquimais, com grande capacidade de formar metástases e geralmente apresentam elevada resistência à terapia. Formam um grupo extremamente heterogêneo com mais de 100 subtipos tumorais, como os sarcomas pleomórficos indiferenciados (SPI) e os sarcomas fusocelulares (SF). Em conjunto, esses dois subgrupos não compartilham semelhanças morfológicas com os outros subtipos. Adicionalmente, carecem de informações moleculares que auxiliem tanto no diagnóstico quanto no prognóstico da doença. O objetivo deste trabalho foi estabelecer e caracterizar modelos pré-clínicos de SPI e SF baseados em Patient-derived xenografts (PDX) e organoids (PDO) para definir uma plataforma de estudos destes tumores e avaliar o potencial de alvos moleculares específicos. Utilizando amostras de 11 pacientes, foi possível estabelecer oito PDX de SPI ou SF. O estabelecimento de organóides de sarcoma se mostrou desafiador para amostras tumorais oriundas diretamente de pacientes ou de PDX e após testarmos diferentes protocolos, conseguimos obter organóides de culturas primárias tumorais. Os organoides e os PDXs gerados foram caracterizados por IHQ e foi possível observar a preservação das características moleculares. Para a caracterização genética, realizamos o sequenciamento de exoma de um conjunto de oito amostras (paciente/PDX/cultura celular) para identificação das principais alterações somáticas. Por fim, padronizamos o ensaio de citotoxicidade à doxorrubicina em três culturas primárias de SPI e SF.

Soft tissue sarcomas (STS) are malignant tumors derived from mesenchymal cells, with a great capacity to form metastases and generally show high resistance to therapy. They form an extremely heterogeneous group with more than 100 tumor subtypes, such as undifferentiated pleomorphic sarcoma (UPS) and fusocellular sarcomas (SS). Taken together, these two subgroups do not share morphological similarities with the other subtypes. Additionally, they lack molecular information that helps both in the diagnosis and prognosis of the disease. The objective of this work was to establish and characterize preclinical models of UPS and SS based on Patient-derived xenografts (PDX) and organoids (PDO) to define a platform for studies of these tumors and to evaluate the potential of specific molecular targets. Using samples from eleven patients, it was possible to establish eight PDX of UPS and SS. The establishment of sarcoma organoids proved to be challenging for tumor samples derived directly from patients or from PDX and after testing different protocols, we were able to obtain organoids from primary tumor cultures. The generated organoids and PDXs were also characterized by IHC and immunofluorescence with sarcoma marker proteins and it was also possible to observe the preservation of molecular characteristics. For genetic characterization, we performed exome sequencing of a set of eight samples (patient/PDX/cell culture) to identify the main somatic alterations. Finally, we standardized the doxorubicin cytotoxicity assay in three primary cultures of SPI and SF.

Vitamin D analogues exhibit antineoplastic activity in breast cancer patient-derived xenograft cells.

Despite advances in breast cancer (BC) treatment, its mortality remains high due to intrinsic or acquired resistance to therapy. Several ongoing efforts are being made to develop novel drugs to treat this pathology with the aim to overcome resistance, prolong patient survival and improve their quality of life. We have previously shown that the non-hypercalcemic vitamin D analogues EM1 and UVB1 display antitumor effects in preclinical studies employing conventional cell lines and animal models developed from these cells. In this work, we explored the antitumor effects of EM1 and UVB1 employing BC cells derived from patient-derived xenografts (PDXs), which are a powerful preclinical tool for testing new drugs. We demonstrated that the analogues reduced the viability of HER2-positive and Triple Negative BC-PDXs. Moreover, using an in vitro model of acquired resistance to Trastuzumab-emtansine, UVB1 displayed anti-proliferative actions under 2D and 3D culture conditions. It inhibited both formation and growth of established organoids. In addition, a direct correlation between UVB1 antitumor effects and VDR expression in PDXs was found. In conclusion, all the results reinforce the potential use of these vitamin D analogues as antitumor agents to treat HER2-positive and Triple Negative BC.

Hippo-YAP1 Is a Prognosis Marker and Potentially Targetable Pathway in Advanced Gallbladder Cancer.

Gallbladder cancer is an aggressive disease with late diagnosis and no efficacious treatment. The Hippo-Yes-associated protein 1 (YAP1) signaling pathway has emerged as a target for the development of new therapeutic interventions in cancers. However, the role of the Hippo-targeted therapy has not been addressed in advanced gallbladder cancer (GBC). This study aimed to evaluate the expression of the major Hippo pathway components mammalian Ste20-like protein kinase 1 (MST1), YAP1 and transcriptional coactivator with PDZ-binding motif (TAZ) and examined the effects of Verteporfin (VP), a small molecular inhibitor of YAP1-TEA domain transcription factor (TEAD) protein interaction, in metastatic GBC cell lines and patient-derived organoids (PDOs). Immunohistochemical analysis revealed that advanced GBC patients had high nuclear expression of YAP1. High nuclear expression of YAP1 was associated with poor survival in GBC patients with subserosal invasion (pT2). Additionally, advanced GBC cases showed reduced expression of MST1 compared to chronic cholecystitis. Both VP treatment and YAP1 siRNA inhibited the migration ability in GBC cell lines. Interestingly, gemcitabine resistant PDOs with high nuclear expression of YAP1 were sensitive to VP treatment. Taken together, our results suggest that key components of the Hippo-YAP1 signaling pathway are dysregulated in advanced gallbladder cancer and reveal that the inhibition YAP1 may be a candidate for targeted therapy.

Anti-tumor activity of the MDM2-TP53 inhibitor BI-907828 in dedifferentiated liposarcoma patient-derived xenograft models harboring MDM2 amplification.

PURPOSE: Dedifferentiated liposarcoma (DDLPS) is a soft tissue malignancy characterized by amplification of the mouse double minute 2 homolog (MDM2) gene. MDM2 is a negative regulator of tumor protein 53 (TP53). We tested the in vivo efficacy of BI-907828, a small molecule inhibitor of the MDM2-TP53 interaction, in two DDLPS patient-derived xenografts (PDX). METHODS: Partially immunodeficient mice were bilaterally engrafted with UZLX-STS3 (n = 24) and UZLX-STS5 (n = 24) human DDLPS tissue harboring MDM2 amplifications. Mice were grouped as follows: (a) vehicle (0.5% hydroxyethylcellullose) 10 ml/kg daily per os (p.o.); (b) doxorubicin 5 mg/kg weekly intraperitoneally (i.p.); (c) BI-907828 2.5 mg/kg daily p.o. and (d) BI-907828 10 mg/kg daily p.o. The treatment lasted for 15 days, all mice treated with BI-907828 were followed for 37 days post-treatment. Efficacy was assessed by tumor volume and histopathological evaluation. RESULTS: The 15-day treatment with 2.5 mg/kg and 10 mg/kg BI-907828 significantly inhibited tumor growth in UZLX-STS5 and -STS3 (p < 0.0001 compared to control for both models). All UZLX-STS5 and -STS3 tumors treated with BI-907828 decreased in size during treatment, and BI-907828-treated UZLX-STS5 tumors even disappeared completely. During the follow-up period, no tumor regrowth was observed in the UZLX-STS5 model and both doses of BI-907828 led to a pathological complete response, whereas a dose-dependent regrowth was seen in the UZLX-STS3 model. CONCLUSION: BI-907828 showed significant anti-tumor activity in DDLPS PDX harboring MDM2 amplifications, providing a strong rationale for early clinical testing of BI-907828 in a DDLPS patient population.

A poor prognosis in human hepatocellular carcinoma is associated with low expression of DPP4

This study aimed to explore the prognostic role of dipeptidyl peptidase 4 (DPP4) expression in hepatocellular carcinoma (HCC). DPP4 expression was measured in formalin-fixed paraffin-embedded specimens that were gathered from 327 HCC patients. Immunohistochemistry analyses were utilized to examine DPP4 expression characteristics and prognostic values (overall survival (OS) and time to recurrence) of DDP4 in HCC tissues. In addition, a patient-derived xenograft (PDX) model was used to assess the correlation between DPP4 expression and tumor growth in vivo. DPP4 was expressed in low levels in HCC tissues in contrast to paired peritumoral tissues (38 cases were down-regulated in a total of 59 cases, 64.4%. P=0.0202). DPP4 expression was significantly correlated with TNM stage (P=0.038), tumor number (P=0.035), and vascular invasion (P=0.024), and significantly reduced in patients who were in TNM stages II and III-V, with multiple tumors, and with microvascular invasion compared to patients with TNM stage I, single tumor, and no microvascular invasion. Notably, HCC tissues with low expression of DPP4 had poor OS (P=0.016) compared with HCC tissues with high expression of DPP4, and results from PDX model showed that tumor growth was significantly faster in HCC patients that lowly expressed DPP4 compared to those with highly expressed DPP4. Our findings suggested that low levels of DPP4 could impact the aggressiveness of HCC and contribute to a poor prognosis.

Pilot Study of an Integrative New Tool for Studying Clinical Outcome Discrimination in Acute Leukemia.

Acute leukemia is a heterogeneous set of diseases affecting children and adults. Current prognostic factors are not accurate predictors of the clinical outcome of adult patients and the stratification of risk groups remains insufficient. For that reason, this study proposes a multifactorial analysis which integrates clinical parameters, ex vivo tumor characterization and behavioral in vivo analysis in zebrafish. This model represents a new approach to understand leukemic primary cells behavior and features associated with aggressiveness and metastatic potential. Xenotransplantation of primary samples from patients newly diagnosed with acute leukemia in zebrafish embryos at 48 hpf was used to asses survival rate, dissemination pattern, and metastatic potential. Seven samples from young adults classified in adverse, favorable or intermediate risk group were characterized. Tumor heterogeneity defined by Leukemic stem cell (LSC) proportion, was performed by metabolic and cell membrane biomarkers characterization. Thus, our work combines all these parameters with a robust quantification strategy that provides important information about leukemia biology, their relationship with specific niches and the existent inter and intra-tumor heterogeneity in acute leukemia. In regard to prognostic factors, leukemic stem cell proportion and Patient-derived xenografts (PDX) migration into zebrafish were the variables with highest weights for the prediction analysis. Higher ALDH activity, less differentiated cells and a broader and random migration pattern are related with worse clinical outcome after induction chemotherapy. This model also recapitulates multiple aspects of human acute leukemia and therefore is a promising tool to be employed not only for preclinical studies but also supposes a new tool with a higher resolution compared to traditional methods for an accurate stratification of patients into worse or favorable clinical outcome.

Establishment of patient-derived tumor xenograft (PDTX) models using samples from CT-guided percutaneous biopsy

This study aimed to investigate the feasibility of the establishment of a human cancer xenograft model using samples from computed tomography (CT)-guided percutaneous biopsy. Fresh tumor tissues obtained from 10 cancer patients by CT-guided percutaneous biopsy were subcutaneously inoculated into NOD-Prkdcem26Il2rgem26Nju (NCG) mice to establish human patient-derived tumor xenograft (PDTX) models. The formation of first and second generation xenografts was observed, and tumor volume was recorded over time. Tumor tissue consistency between the PDTX model and primary tumors in patients was compared using H&E staining and immunohistochemistry. Pharmacodynamic tests of clinically used chemotherapeutic drugs were conducted on second generation xenografts, and their effects on tumor growth and body weight were observed. CT-guided percutaneous biopsy samples were successfully collected from 10 patients with advanced cancers. The PDTX model was established in mice using tumor samples obtained from 4 cancer patients, including one small cell carcinoma sample, two adenocarcinoma samples, and one squamous cell carcinoma sample. The success rate was 40%. The obtained PDTX model maintained a degree of differentiation, and morphological and structural characteristics were similar to primary tumors. The pharmacodynamic test of chemotherapeutic drugs in the PDTX model revealed a therapeutic effect on tumor growth, as expected. CT-guided percutaneous biopsy samples can be effectively used to establish a PDTX model, and test these chemotherapy regimens.

The combinational therapy of trastuzumab and cetuximab inhibits tumor growth in a patient-derived tumor xenograft model of gastric cancer.

PURPOSE: Gastric cancer (GC) is one of the leading causes of cancer mortality worldwide. Although therapeutic strategies for GC have improved, the prognosis for advanced GC remains poor. Herein, the present study sought to design a personalized cancer therapy specific to a stage III GC patient. METHODS: The tumor was surgically removed and was used to establish a patient-derived tumor xenograft (PDTX) model utilizing nude mice. Various molecular-targeted anticancer treatments were tested in the study, including control (no treatment), bevacizumab, cetuximab, bevacizumab + cetuximab, trastuzumab, and trastuzumab + cetuximab. RESULTS: Trastuzumab + cetuximab treatment exhibited the best antitumor growth effect, followed by trastuzumab, bevacizumab + cetuximab, cetuximab, and bevacizumab. Similarly, trastuzumab + cetuximab was also the most effective treatment at inducing apoptosis and cell cycle arrest in primary cultures of the patient's gastric cancer cells. Among all treatments tested in the study, trastuzumab + cetuximab showed the most profound effect in reducing the protein expression of proliferation and metastatic markers (VEGF, MMP-7, EGFT, Ki-67 and, PCNA) in tumors obtained from PDTX models, which may be the mechanism underlying the profound antitumor growth effect exerted by trastuzumab + cetuximab. CONCLUSIONS: The data indicate that trastuzumab + cetuximab combinational therapy should be the most effective antitumor growth therapy for the GC patient whom we took the cancer cells from.