Search

Anaplastic large-cell lymphoma (ALCL) is a T-cell malignancy predominantly driven by the oncogenic anaplastic lymphoma kinase (ALK), accounting for approximately 15% of all paediatric non-Hodgkin lymphoma. Patients with central nervous system (CNS) relapse are particularly difficult to treat with a 3-year overall survival of 49% and a median survival of 23.5 months. The second-generation ALK inhibitor brigatinib shows superior penetration of the blood-brain barrier unlike the first-generation drug crizotinib and has shown promising results in ALK+ non-small-cell lung cancer. However, the benefits of brigatinib in treating aggressive paediatric ALK+ ALCL are largely unknown. We established a patient-derived xenograft (PDX) resource from ALK+ ALCL patients at or before CNS relapse serving as models to facilitate the development of future therapies. We show in vivo that brigatinib is effective in inducing the remission of PDX models of crizotinib-resistant (ALK C1156Y, TP53 loss) ALCL and furthermore that it is superior to crizotinib as a second-line approach to the treatment of a standard chemotherapy relapsed/refractory ALCL PDX pointing to brigatinib as a future therapeutic option.

Background Hepatocellular carcinoma (HCC) patient‐derived xenograft (PDX) models hold potential to advance knowledge in HCC biology to help improve systemic therapies. Beside hepatitis B virus‐associated tumors, HCC is poorly established in PDX. Methods PDX formation from fresh HCC biopsies were obtained and implanted intrahepatically or in subrenal capsule (SRC). Mouse liver injury was induced in immunodeficient Fah−/− mice through cycling off nitisinone after HCC biopsy implantation, versus continuous nitisinone as non‐liver injury controls. Mice with macroscopically detectable PDX showed rising human alpha1‐antitrypsin (hAAT) serum levels, and conversely, no PDX was observed in mice with undetectable hAAT. Results Using rising hAAT as a marker for PDX formation, 20 PDX were established out of 45 HCC biopsy specimens (44%) reflecting the four major HCC etiologies most commonly identified at Memorial SloanKettering similar to many other institutions in the United States. PDX was established only in severely immunodeficient mice lacking lymphocytes and NK cells. Implantation under the renal capsule improved PDX formation two‐fold compared to intrahepatic implantation. Two out of 18 biopsies required murine liver injury to establish PDX, one associated with hepatitis C virus and one with alcoholic liver disease. PDX tumors were histologically comparable to biopsy specimens and 75% of PDX lines could be passaged. Conclusions Using cycling off nitisinone‐induced liver injury, HCC biopsies implanted under the renal capsule of severely immunodeficient mice formed PDX with 57% efficiency as determined by rising hAAT levels. These findings facilitate a more efficient make‐up of PDX for research into subset‐specific HCC.

Myxoid liposarcoma (MLPS) is a rare soft-tissue sarcoma characterised by the expression of FUS-DDIT3 chimera. Trabectedin has shown significant clinical anti-tumour activity against MLPS. To characterise the molecular mechanism of trabectedin sensitivity and of resistance against it, we integrated genomic and transcriptomic data from treated mice bearing ML017 or ML017/ET, two patient-derived MLPS xenograft models, sensitive to and resistant against trabectedin, respectively. Longitudinal RNA-Seq analysis of ML017 showed that trabectedin acts mainly as a transcriptional regulator: 15 days after the third dose trabectedin modulates the transcription of 4883 genes involved in processes that sustain adipocyte differentiation. No such differences were observed in ML017/ET. Genomic analysis showed that prolonged treatment causes losses in 4p15.2, 4p16.3 and 17q21.3 cytobands leading to acquired-resistance against the drug. The results dissect the complex mechanism of action of trabectedin and provide the basis for novel combinatorial approaches for the treatment of MLPS that could overcome drug-resistance.

Acute lymphoblastic leukemia (ALL) represents the most frequent malignancy in children, and relapse/refractory (r/r) disease is difficult to treat, both in children and adults. In search for novel treatment options against r/r ALL, we studied inhibitor of apoptosis proteins (IAP) and Smac mimetics (SM). SM-sensitized r/r ALL cells towards conventional chemotherapy, even upon resistance against SM alone. The combination of SM and chemotherapy-induced cell death via caspases and PARP, but independent from cIAP-1/2, RIPK1, TNFα or NF-κB. Instead, XIAP was identified to mediate SM effects. Molecular manipulation of XIAP invivo using microRNA-30 flanked shRNA expression in cell lines and patient-derived xenograft (PDX) models of r/r ALL mimicked SM effects and intermediate XIAP knockdown-sensitized r/r ALL cells towards chemotherapy-induced apoptosis. Interestingly, upon strong XIAP knockdown, PDX r/r ALL cells were outcompeted invivo, even in the absence of chemotherapy. Our results indicate a yet unknown essential function of XIAP in r/r ALL and reveal XIAP as a promising therapeutic target for r/r ALL.

Triple-negative breast cancers (TNBCs) currently have limited treatment options; however, PD-L1 is an indicator of susceptibility to immunotherapy. Currently, assessment of PD-L1 is limited to biopsy samples. These limitations may be overcome with molecular imaging. In this work, we describe chemistry development and optimization, in vitro, in vivo, and dosimetry of [89Zr]-Atezolizumab for PD-L1 imaging. Atezolizumab was conjugated to DFO and radiolabeled with 89Zr. Tumor uptake and heterogeneity in TNBC xenograft and patient-derived xenograft (PDX) mouse models were quantified following [89Zr]-Atezolizumab-PET imaging. PD-L1 expression in TNBC PDX models undergoing therapy and immunohistochemistry (IHC) was used to validate imaging. SUV from PET imaging was quantified and used to identify heterogeneity. PET/CT imaging using [89Zr]-Atezolizumab identified a significant increase in tumor:muscle SUVmean 1 and 4 days after niraparib therapy and revealed an increased trend in PD-L1 expression following other cytotoxic therapies. A preliminary dosimetry study indicated the organs that will receive a higher dose are the spleen, adrenals, kidneys, and liver. [89Zr]-Atezolizumab PET/CT imaging reveals potential for the noninvasive detection of PD-L1-positive TNBC tumors and allows for quantitative and longitudinal assessment. This has potential significance for understanding tumor heterogeneity and monitoring early expression changes in PD-L1 induced by therapy.

Background: The application of chimeric antigen receptor (CAR) NK cells in solid tumors is hindered by lack of tumor-specific targets and inefficient CAR NK cell efficacy. It has been reported that mesothelin (MSLN) may be an ideal immunotherapy target for gastric cancer. However, the feasibility of using anti-MSLN CAR NK cells to treat gastric cancer remains to be studied. Methods: MSLN expression in primary human gastric cancer, normal tissues and cell lines were detected. MSLN and CD19 targeted CAR NK-92 (MSLN- and CD19-CAR NK) cells were constructed, purified and verified. N87, MKN-28, AGS and Huh-7 cells expressing the GFP and luciferase genes were transduced. Cell- and patient-derived xenograft (PDX) were established via NSG mice. The ability of MSLN-CAR NK cells to kill MSLN-positive gastric cancer cells were evaluated in vitro and in vivo. Results: MSLN-CAR NK cells can specifically kill MSLN-positive gastric cancer cells (N87, MKN-28 and AGS), rather than MSLN negative cell (Huh-7), in vitro. Moreover, compared with parental NK-92 cells and CD19-CAR NK cells, stronger cytokine secretions were secreted in MSLN-CAR NK cells cocultured with N87, MKN-28 and AGS. Furthermore, MSLN-CAR NK cells can effectively eliminate gastric cancer cells in both subcutaneous and intraperitoneal tumor models. They could also significantly prolong the survival of intraperitoneally tumor-bearing mice. More importantly, the potent antitumor effect and considerable NK cell infiltration were observed in the patient-derived xenograft treated with MSLN-CAR NK cells, which further warranted the therapeutic effects of MSLN-CAR NK cells to treat gastric cancer. Conclusion: These results demonstrate that MSLN-CAR NK cells possess strong antitumor activity and represent a promising therapeutic approach to gastric cancer.

Targeting the androgen receptor (AR) signalling pathway remains the main therapeutic option for advanced prostate cancer. However, resistance to AR‐targeting inhibitors represents a great challenge, highlighting the need for new therapies. Activation of the PI3K/AKT pathway and increased expression of histone deacetylases (HDACs) are common aberrations in prostate cancer, suggesting that inhibition of such targets may be a viable therapeutic strategy for this patient population. Previous reports demonstrated that combination of PI3K inhibitors (PI3KIs) with histone deacetylase inhibitors (HDACIs) resulted in synergistic antitumour activities against preclinical models of prostate cancer. In this study, we demonstrate that the novel dual PI3K and HDAC inhibitor CUDC‐907 has promising antitumour activity against prostate cancer cell lines in vitro and castration‐resistant LuCaP 35CR patient‐derived xenograft (PDX) mouse model in vivo. CUDC‐907‐induced apoptosis was partially dependent on Mcl‐1, Bcl‐xL, Bim and c‐Myc. Further, down‐regulation of Wee1, CHK1, RRM1 and RRM2 contributed to CUDC‐907‐induced DNA damage and apoptosis. In the LuCaP 35CR PDX model, treatment with CUDC‐907 resulted in significant inhibition of tumour growth. These findings support the clinical development of CUDC‐907 for the treatment of prostate cancer.

There are three subtypes of undifferentiated human conventional osteosarcoma (HCOS): osteoblastic osteosarcoma (OOS), chondroblastic osteosarcoma (COS), and fibroblastic osteosarcoma (FOS). HCOS also exhibits heterogeneous pathological maldifferentiation in individual patients. Currently, the mechanism regulating HCOS differentiation remains unclear, and therapies are ineffective. Osteopontin (OPN) and osteocalcin (OCN) are markers of osteoblast maturation, and their expression is inhibited in HCOS. A previous study found that PLK2 inhibited TAp73 phosphorylation and consequent anti‐OS function of TAp73 in OS cells with enriched TAp73. TAp73 was also reported to regulate bone cell calcification. Here, OOS was found to have higher TAp73 levels and PLK2 expression than those in COS, which is correlated with HCOS maldifferentiation according to Spearman analysis and affects patient prognosis according to Kaplan‐Meier survival analysis. In the conventional OS cell‐line Saos2 and in patient‐derived xenograft OS (PDX‐OS) cells, increased PLK2 expression owing to abundant TAp73 levels affected OPN and OCN content as measured by RT‐PCR and Western blotting, and alizarin red staining showed that PLK2 affected calcium deposition in OS cells. In addition, PLK2 inhibition in PDX‐OS cells prohibited clone formation, as indicated by a clonogenic assay, and sensitized OS cells to cisplatin (CDDP) (which consequently limited proliferation), as shown by the CCK‐8 assay. In an established PDX animal model with abundant TAp73 levels, PLK2 inhibition or CDDP treatment prevented tumor growth and prolonged median survival. The combined therapeutic effect of PLK2 inhibition with CDDP treatment was better than that of either monotherapy. These results indicate that increased PLK2 levels due to enriched TAp73 affect osteogenic differentiation and maturation and OS prognosis. In conclusion, PLK2 is a potential target for differentiation therapy of OS with enriched TAp73., This study indicates that PLK2 modulation of highly abundant TAp73 to regulate the clonogenicity, proliferation, osteogenic differentiation, and maturation of OS cells. In addition, PLK2 is a potential therapeutic target in preclinical PDX hosts that can sensitize tumors to the antitumor effect of CDDP and prolong the medium survival of the hosts. In conclusion, PLK2 may be an ideal target for OS differentiation therapy.

Background Cancer cachexia is a life‐threatening metabolic syndrome that causes significant loss of skeletal muscle mass and significantly increases mortality in cancer patients. Currently, there is an urgent need for better understanding of the molecular pathophysiology of this disease so that effective therapies can be developed. The majority of pre‐clinical studies evaluating skeletal muscle's response to cancer have focused on one or two pre‐clinical models, and almost all have focused specifically on limb muscles. In the current study, we reveal key differences in the histology and transcriptomic signatures of a limb muscle and a respiratory muscle in orthotopic pancreatic cancer patient‐derived xenograft (PDX) mice. Methods To create four cohorts of PDX mice evaluated in this study, tumours resected from four pancreatic ductal adenocarcinoma patients were portioned and attached to the pancreas of immunodeficient NSG mice. Results Body weight, muscle mass, and fat mass were significantly decreased in each PDX line. Histological assessment of cryosections taken from the tibialis anterior (TA) and diaphragm (DIA) revealed differential effects of tumour burden on their morphology. Subsequent genome‐wide microarray analysis on TA and DIA also revealed key differences between their transcriptomes in response to cancer. Genes up‐regulated in the DIA were enriched for extracellular matrix protein‐encoding genes and genes related to the inflammatory response, while down‐regulated genes were enriched for mitochondria related protein‐encoding genes. Conversely, the TA showed up‐regulation of canonical atrophy‐associated pathways such as ubiquitin‐mediated protein degradation and apoptosis, and down‐regulation of genes encoding extracellular matrix proteins. Conclusions These data suggest that distinct biological processes may account for wasting in different skeletal muscles in response to the same tumour burden. Further investigation into these differences will be critical for the future development of effective clinical strategies to counter cancer cachexia.

Therapeutic RNAs, such as antisense oligonucleotides (ASOs), aptamers, small-interfering RNAs (siRNAs), microRNAs (miRs or miRNAs), messenger RNAs (mRNAs), and guide RNAs (gRNAs), represent a novel class of modalities that not only increase the molecular diversity of medications but also expand the range of druggable targets. To develop noncoding RNA therapeutics for the treatment of cancer diseases, we have established a novel robust RNA bioengineering platform to achieve high-yield and large-scale production of true biologic RNA agents, which are proven to be functional in the control of target gene expression and effective in the management of tumor progression in various models. Herein, we describe the methods for bioengineered RNA (BioRNA or BERA) therapy in patient-derived organoids (PDOs) in vitro and patient-derived xenograft (PDX) mouse models in vivo. The efficacy of a BioRNA, miR-1291, in the inhibition of pancreatic cancer PDO and PDX growth is exemplified in this chapter.

El cáncer sigue siendo una de las principales causas de muerte en el mundo. El glioblastoma multiforme (GBM) es el tumor más agresivo y prevalente del sistema nervioso central, con una de supervivencia de tan solo 15 a 18 meses desde su diagnóstico. Pese a que se ha observado que su incidencia varía en relación con factores como la edad, la raza o el sexo, el GBM puede desarrollarse en cualquier persona. Además, la efectividad de los tratamientos tradicionales está condicionada por factores como el microambiente tumoral y el subtipo molecular de GBM que se desarrolle, siendo necesaria la búsqueda de nuevas terapias dirigidas y específicas a cada paciente. Por estos motivos, la presente investigación se centró en el desarrollo de un nuevo modelo preclínico para el estudio del GBM basado en embriones de pollo ex ovo. En base a estudios previos de este grupo de investigación, se seleccionó la línea celular de GBM humana GB-39, para la generación de un xenoinjerto derivado de paciente (PDX) en el cerebro de los embriones de pollo. Los embriones manifestaron un desarrollo morfológico normal en el tiempo que duraron los experimentos y los tumores generados en ellos fueron capaces de injertarse y proliferar, recapitulando las características de los tumores de GBM humanos. Además, el análisis de los marcadores de células microgliales Iba1 y CD11B sí confirmaron una activación moderada de la microglía que se correspondía con la progresión normal del tumor. Los análisis con los marcadores específicos GLAST y CD44, revelaron que los tumores no presentaban patrones de crecimiento invasivos ni infiltración con transición epitelio-mesénquima que pudiera derivar en metástasis. Por último, se llevó a cabo una prueba de concepto, que consistió en la inyección e integración de exosomas, para comprobar el potencial del modelo en el estudio de la vehiculiazación de fármacos. Los datos recogidos en este proyecto confirman que el modelo desarrollado puede ser usado como modelo preclínico para testar in vivo tratamientos contra el GBM humano, pudiendo utilizarse en futuros estudios experimentales en el campo de la biomedicina. Cancer remains one of the leading causes of death in the world. Glioblastoma multiforme (GBM) is the most aggressive and prevalent tumor of the central nervous system, reaching a survival period of only 15 to 18 months after diagnosis. Although it has been observed that its incidence varies in relation to factors such as age, race or gender, GBM can be developed by any person. Furthermore, the effectiveness of traditional treatments is conditioned by factors such as the tumor microenvironment and the molecular subtype of GBM, making necessary the search for new targeted therapies specific to each patient. For these reasons, the present research focused on the development of a new preclinical model to study of GBM that is based on chicken embryos ex ovo. According to previous studies of this research group, the GBM human derived cell line GB-39 was selected for the generation of a patient-derived xenograft (PDX) in the brain of chicken embryos. In this study, we observed that the embryos showed a normal morphological development during all experimental process. It was also possible to confirm, by histological and immunohistochemical analysis of the markers KI67, GFAP and S100B, that the PDX-generated tumors were able to engraft and proliferate, recapitulating the characteristics of human GBM tumors. In addition, analysis with GLAST and CD44 specific markers revealed that the tumors did not exhibit invasive growth patterns or infiltration with epithelial-mesenchymal transition that could lead to metastasis. However, analysis of the microglial cell markers Iba1 and CD11B did confirm moderate microglial activation consistent with normal tumor progression. Finally, a proof of concept, consisting of the injection and integration of exosomes, was carried out to test the potential of the model in the study of drug delivery. The data collected in this project confirm that the developed model can be used as a preclinical model to test in vivo human GBM treatments, which will contribute to future experimental studies in the field of biomedicine.

Background Accurate predictions of tumor dissemination risks and medical treatment outcomes are critical to personalize therapy. Patient-derived xenograft (PDX) models in mice have demonstrated high accuracy in predicting therapeutic outcomes, but methods for predicting tumor invasiveness and early stages of vascular/lymphatic dissemination are still lacking. Here we show that a zebrafish tumor xenograft (ZTX) platform based on implantation of PDX tissue fragments recapitulate both treatment outcome and tumor invasiveness/dissemination in patients, within an assay time of only 3 days. Methods Using a panel of 39 non-small cell lung cancer PDX models, we developed a combined mouse-zebrafish PDX platform based on direct implantation of cryopreserved PDX tissue fragments into zebrafish embryos, without the need for pre-culturing or expansion. Clinical proof-of-principle was established by direct implantation of tumor samples from four patients. Results The resulting ZTX models responded to Erlotinib and Paclitaxel, with similar potency as in mouse-PDX models and the patients themselves, and resistant tumors similarly failed to respond to these drugs in the ZTX system. Drug response was coupled to elevated expression of EGFR, Mdm2, Ptch1 and Tsc1 (Erlotinib), or Nras and Ptch1 (Paclitaxel) and reduced expression of Egfr, Erbb2 and Foxa (Paclitaxel). Importantly, ZTX models retained the invasive phenotypes of the tumors and predicted lymph node involvement of the patients with 91% sensitivity and 62% specificity, which was superior to clinically used tests. The biopsies from all four patient tested implanted successfully, and treatment outcome and dissemination were quantified for all patients in only 3 days. Conclusions We conclude that the ZTX platform provide a fast, accurate, and clinically relevant system for evaluation of treatment outcome and invasion/dissemination of PDX models, providing an attractive platform for combined mouse-zebrafish PDX trials and personalized medicine.

Background Graft-contaminating tumor cells correlate with inferior outcome in high-risk neuroblastoma patients undergoing hematopoietic stem cell transplantation and can contribute to relapse. Motivated by the potential therapeutic benefit of tumor cell removal as well as the high prognostic and diagnostic value of isolated circulating tumor cells from stem cell grafts, we established a label-free acoustophoresis-based microfluidic technology for neuroblastoma enrichment and removal from peripheral blood progenitor cell (PBPC) products. Methods Neuroblastoma patient-derived xenograft (PDX) cells were spiked into PBPC apheresis samples as a clinically relevant model system. Cells were separated by ultrasound in an acoustophoresis microchip and analyzed for recovery, purity and function using flow cytometry, quantitative real-time PCR and cell culture. Results PDX cells and PBPCs showed distinct size distributions, which is an important parameter for efficient acoustic separation. Acoustic cell separation did not affect neuroblastoma cell growth. Acoustophoresis allowed to effectively separate PDX cells from spiked PBPC products. When PBPCs were spiked with 10% neuroblastoma cells, recoveries of up to 98% were achieved for PDX cells while more than 90% of CD34+ stem and progenitor cells were retained in the graft. At clinically relevant tumor cell contamination rates (0.1 and 0.01% PDX cells in PBPCs), neuroblastoma cells were depleted by more than 2-log as indicated by RT-PCR analysis of PHOX2B, TH and DDC genes, while > 85% of CD34+ cells could be retained in the graft. Conclusion These results demonstrate the potential use of label-free acoustophoresis for PBPC processing and its potential to develop label-free, non-contact tumor cell enrichment and purging procedures for future clinical use.

Background Patient-derived xenografts (PDX) mice models play an important role in preclinical trials and personalized medicine. Sharing data on the models is highly valuable for numerous reasons – ethical, economical, research cross validation etc. The EurOPDX Consortium was established 8 years ago to share such information and avoid duplicating efforts in developing new PDX mice models and unify approaches to support preclinical research. EurOPDX Data Portal is the unified data sharing platform adopted by the Consortium. Main body In this paper we describe the main features of the EurOPDX Data Portal (https://dataportal.europdx.eu/), its architecture and possible utilization by researchers who look for PDX mice models for their research. The Portal offers a catalogue of European models accessible on a cooperative basis. The models are searchable by metadata, and a detailed view provides molecular profiles (gene expression, mutation, copy number alteration) and treatment studies. The Portal displays the data in multiple tools (PDX Finder, cBioPortal, and GenomeCruzer in future), which are populated from a common database displaying strictly mutually consistent views. (Short) Conclusion EurOPDX Data Portal is an entry point to the EurOPDX Research Infrastructure offering PDX mice models for collaborative research, (meta)data describing their features and deep molecular data analysis according to users’ interests.

Simple Summary Our findings indicated that the EGF-EGFR pathway was highly activated in very young patients with recurrent or metastatic NPC. High EGFR expression in patients with metastatic NPC resulted in poor clinical outcomes. To examine whether the EGFR pathway serves as a therapeutic target for NPC, NPC patient-derived xenograft (PDX) and NPC cell lines were treated with EGFR inhibitors (EGFRi) and a cell cycle blocker. Either EGFRi or cell cycle blocker treatment alone could reduce NPC cell growth and PDX tumor growth. Furthermore, combination treatment exerted an additive suppression effect on PDX tumor growth. This study provides promising evidence that EGFRi used in combination with a cell cycle blocker may be used to treat patients with NPC. Abstract Background: Nasopharyngeal carcinoma (NPC) involves host genetics, environmental and viral factors. In clinical observations, patients of young and old ages were found to have higher recurrence and metastatic rates. Methods: Cytokine array was employed to screen druggable target(s). The candidate target(s) were confirmed through patient-derived xenografts (PDXs) and a new EBV-positive cell line, NPC-B13. Results: Overexpression of epithelial growth factor (EGF) and EGF receptor (EGFR) was detected in young patients than in older patients. The growth of NPC PDX tumors and cell lines was inhibited by EGFR inhibitors (EGFRi) cetuximab and afatinib when used separately or in combination with the cell cycle blocker palbociclib. Western blot analysis of these drug-treated PDXs demonstrated that the blockade of the EGF signaling pathway was associated with a decrease in the p-EGFR level and reduction in PDX tumor size. RNA sequencing results of PDX tumors elucidated that cell cycle-related pathways were suppressed in response to drug treatments. High EGFR expression (IHC score ≥ grade 3) was correlated with poor survival in metastatic patients (p = 0.008). Conclusions: Our results provide encouraging preliminary data related to the combination treatment of EGFRi and palbociclib in patients with NPC.

Summary Acute lymphoblastic leukemia (ALL) dissemination to the central nervous system (CNS) is a challenging clinical problem whose underlying mechanisms are poorly understood. Here, we show that primary human ALL samples injected into the femora of immunodeficient mice migrate to the skull and vertebral bone marrow and provoke bone lesions that enable passage into the subarachnoid space. Treatment of leukemia xenografted mice with a biologic antagonist of receptor activator of nuclear factor κB ligand (RANKL) blocks this entry route. In addition to erosion of cranial and vertebral bone, samples from individuals with B-ALL also penetrate the blood-cerebrospinal fluid barrier of recipient mice. Co-administration of C-X-C chemokine receptor 4 (CXCR4) and RANKL antagonists attenuate both identified routes of entry. Our findings suggest that targeted RANKL and CXCR4 pathway inhibitors could attenuate routes of leukemia blast CNS invasion and provide benefit for B-ALL-affected individuals., Graphical abstract, Highlights • Primary human B-ALL cells use multiple independent transit mechanisms to invade the CNS • B-ALL migrate to skull or vertebral bone marrow in patient-derived xenograft (PDX) mice • B-ALL blasts provoke bone lesions through which they enter the subarachnoid space • RANKL and CXCR4 antagonists attenuate B-ALL entry into CNS in PDX mice, Rajakumar et al. show that inhibitors targeting immune receptors attenuate primary human B cell leukemia (B-ALL) cells transplanted into mice from invading the central nervous system (CNS). The findings in this study support clinical efforts to block these specific mechanisms of CNS invasion in B-ALL-affected individuals with targeted therapies.

KRAS‒PDEδ interaction is revealed as a promising target for suppressing the function of mutant KRAS. The bottleneck in clinical development of PDEδ inhibitors is the poor antitumor activity of known chemotypes. Here, we identified novel spiro-cyclic PDEδ inhibitors with potent antitumor activity both in vitro and in vivo. In particular, compound 36l (KD = 127 ± 16 nmol/L) effectively bound to PDEδ and interfered with KRAS–PDEδ interaction. It influenced the distribution of KRAS in Mia PaCa-2 cells, downregulated the phosphorylation of t-ERK and t-AKT and promoted apoptosis of the cells. The novel inhibitor 36l exhibited significant in vivo antitumor potency in pancreatic cancer patient-derived xenograft (PDX) models. It represents a promising lead compound for investigating the druggability of KRAS‒PDEδ interaction., Graphical abstract Compound 36l is a novel spiro-cyclic PDEδ inhibitor (KD = 127 ± 16 nmol/L). It inhibited the growth of Mia PaCa-2 cells (IC50 = 6.67 ± 1.7 μmol/L) and exhibited significant in vivo antitumor potency in pancreatic cancer patient-derived xenograft (PDX) models.Image 1

Simple Summary Prostate cancer is one of the most frequently diagnosed cancers in men and is the second leading cause of cancer death in developed countries. Current therapeutics that target the androgen receptor (AR) are only transiently effective. Anti-AR therapy-resistant tumors often emerge with vast cellular and molecular alterations and present themselves at the clinic in more deadly forms including metastatic castration-resistant prostate cancer (mCRPC) or neuroendocrine prostate cancer (NEPC). One emerging strategy in effective treatment of the advanced forms of prostate cancer is to target drivers other than AR. The present study shows that the nuclear receptor RORγ and the serine/threonine kinase PBK form a regulatory loop in hyperactive AR signaling. It also demonstrates that orally administered, small-molecule antagonists/inverse agonists of RORγ are effective in blocking the growth of the mCRPC tumors. Our findings provide a rationale for therapeutic targeting of RORγ alone or in combination with PBK inhibitors for the advanced forms of prostate cancer. Abstract Metastatic castration-resistant prostate cancer (mCRPC) is a highly aggressive disease with few therapeutic options. Hyperactive androgen receptor (AR) signaling plays a key role in CRPC progression. Previously, we identified RAR-related orphan receptor gamma (RORγ) as a novel key driver of AR gene overexpression and increased AR signaling. We report here that several RORγ antagonists/inverse agonists including XY018 and compound 31 were orally effective in potent inhibition of the growth of tumor models including patient-derived xenograft (PDX) tumors. RORγ controls the expression of multiple aggressive-tumor gene programs including those of epithelial-mesenchymal transition (EMT) and invasion. We found that PDZ binding kinase (PBK), a serine/threonine kinase, is a downstream target of RORγ that exerts the cellular effects. Alterations of RORγ expression or function significantly downregulated the mRNA and protein level of PBK. Our further analyses demonstrated that elevated PBK associates with and stabilizes RORγ and AR proteins, thus constituting novel, interlocked feed-forward loops in hyperactive AR and RORγ signaling. Indeed, dual inhibition of RORγ and PBK synergistically inhibited the expression and function of RORγ, AR, and AR-V7, and the growth and survival of CRPC cells. Therefore, our study provided a promising, new strategy for treatment of advanced forms of prostate cancer.

Simple Summary Advances have been made in the study of NSCLC tumors using in vivo models, such as patient-derived xenografts (PDXs). However, the number of PDX models that can represent the heterogeneity of NSCLC between different individuals is still limited. We successfully established nine PDX mice, from which lung adenocarcinoma tumors bearing the KRAS-G12C mutation were the most frequently grafted. We show that the most aggressive tumors have a greater implantation capacity, and the success of their implantation is indicative of a poor prognosis. By using H-score to quantify cell proliferation and mesenchymal markers, we show that PDX tumors evolved towards a more proliferative and mesenchymal phenotype associated with higher protein levels of Ki67, vimentin, and ezrin, suggesting that the evaluation of their combined expression could be used as a prognostic marker to study disease progression. These PDX models provide a valuable platform for NSCLC translational research. Abstract Non-small-cell lung cancer (NSCLC) is the leading cause of cancer death worldwide. The high mortality is very often a consequence of its late diagnosis when the cancer is already locally advanced or has disseminated. Advances in the study of NSCLC tumors have been achieved by using in vivo models, such as patient-derived xenografts. Apart from drug screening, this approach may also be useful for study of the biology of the tumors. In the present study, surgically resected primary lung cancer samples (n = 33) were implanted in immunodeficient mice, and nine were engrafted successfully, including seven adenocarcinomas, one squamous-cell carcinoma, and one large-cell carcinoma. ADC tumors bearing the KRAS-G12C mutation were the most frequently engrafted in our PDX collection. Protein expression of vimentin, ezrin, and Ki67 were evaluated in NSCLC primary tumors and during serial transplantation by immunohistochemistry, using H-score. Our data indicated a more suitable environment for solid adenocarcinoma, compared to other lung tumor subtypes, to grow and preserve its architecture in mice, and a correlation between higher vimentin and ezrin expression in solid adenocarcinomas. A correlation between high vimentin expression and lung adenocarcinoma tumors bearing KRAS-G12C mutation was also observed. In addition, tumor evolution towards more proliferative and mesenchymal phenotypes was already observed in early PDX tumor passages. These PDX models provide a valuable platform for biomarker discovery and drug screening against tumor growth and EMT for lung cancer translational research.

Background Dedifferentiated liposarcoma (DDLPS), a tumor that lacks effective treatment strategies and is associated with poor outcomes, expresses amplified MDM2 in the presence of wild-type p53. MDM2 ubiquitination of p53 facilitates its XPO1-mediated nuclear export, thus limiting p53 tumor suppressor functions. Consequently, nuclear export is a rational target in DDLPS. We directly compared the antitumor activity of the first-in class XPO1 inhibitor selinexor and doxorubicin, the standard front-line therapy in sarcomas, in DDLPS patient-derived xenografts (PDXs) and primary cell lines. Methods Drug activity was assessed in three PDXs (and two corresponding cell lines) established from the dedifferentiated component of primary untreated retroperitoneal DDLPS with myogenic (N = 2) and rhabdomyoblastic (N = 1) differentiation from patients who underwent surgery. These models were marked by amplification of MDM2, CDK4 and HMGA2 genes. Results Selinexor was moderately active in the three PDXs but achieved greater tumor response compared to doxorubicin (maximum tumor volume inhibition: 46–80 % vs. 37–60 %). The PDX harboring rhabdomyoblastic dedifferentiation showed the highest sensitivity to both agents. PDX response to selinexor and doxorubicin was not associated with the extent of MDM2 and CDK4 gene amplification. Interestingly, the most chemosensitive PDX model showed the lowest extent of HMGA2 amplification. Selinexor was also more efficient than doxorubicinin in inducing an apoptotic response in PDXs and cell lines. Consistently, an increased nuclear accumulation of p53 was seen in all selinexor-treated models. In addition, a time-dependent decrease of survivin expression, with an almost complete abrogation of the cytoplasmic anti-apoptotic pool of this protein, was observed as a consequence of the decreased acetylation/activation of STAT3 and the increased ubiquitination of nuclear survivin. Conclusions Selinexor showed a moderate antitumor activity in three DDLPS PDXs, which was, however, consistently higher than doxorubicin across all different models regardless the extent of MDM2 amplification and the histological differentiation. The depletion of survivin protein seems to significantly contribute to the induction of apoptosis through which selinexor exerts its antitumor activity.

[This corrects the article .].

Cancer patient-derived xenografts (PDXs) better preserve tumor characteristics and microenvironment than traditional cancer cell line derived xenografts and are becoming a valuable model in translational cancer research and personalized medicine. We have established a PDX model for colorectal cancer (CRC) in CIEA NOG mice with a 50% engraftment rate. Tumor fragments from patients with CRC (n = 5) were engrafted in four mice per tumor (n = 20). Mice with established PDXs received a liquid diet enriched with fish oil or placebo, and fatty acid profiling was performed to measure fatty acid content in whole blood. Moreover, a biobank consisting of tissue and blood samples from patients was established. Histology, immunohistochemistry and in situ hybridization procedures were used for staining of tumor and xenograft tissue slides. Results demonstrate that key histological characteristics of the patients’ tumors were retained in the established PDXs, and the liquid diets were consumed as intended by the mice. Some of the older mice developed lymphomas that originated from human Ki67+, CD45+, and EBV+ lymphoid cells. We present a detailed description of the process and methodology, as well as possible issues that may arise, to refine the method and improve PDX engraftment rate for future studies. The established PDX model for CRC can be used for exploring different cancer treatment regimes, and liquid diets enriched with fish oil may be successfully delivered to the mice through the drinking flasks.

An in vitro assay system using patient-derived tumor models represents a promising preclinical cancer model that replicates the disease better than traditional cell culture models. Patient-derived tumor organoid (PDO) and patient-derived tumor xenograft (PDX) models have been previously established from different types of human tumors to recapitulate accurately and efficiently their tissue architecture and function. However, these models have low throughput and are challenging to construct. Thus, the present study aimed to establish a simple in vitro high-throughput assay system using PDO and PDX models. Furthermore, the current study aimed to evaluate different classes of anticancer drugs, including chemotherapeutic, molecular targeted and antibody drugs, using PDO and PDX models. First, an in vitro high-throughput assay system was constructed using PDO and PDX established from solid and hematopoietic tumors cultured in 384-well plates to evaluate anticancer agents. In addition, an in vitro evaluation system of the immune response was developed using PDO and PDX. Novel cancer immunotherapeutic agents with marked efficacy have been used against various types of tumor. Thus, there is an urgent need for in vitro functional potency assays that can simulate the complex interaction of immune cells with tumor cells and can rapidly test the efficacy of different immunotherapies or antibody drugs. An evaluation system for the antibody-dependent cellular cytotoxic activity of anti-epidermal growth factor receptor antibody and the cytotoxic activity of activated lymphocytes, such as cytotoxic T lymphocytes and natural killer cells, was constructed. Moreover, immune response assay systems with bispecific T-cell engagers were developed using effector cells. The present results demonstrated that in vitro assay systems using PDO and PDX may be suitable for evaluating anticancer agents and immunotherapy potency with high reproducibility and simplicity.

Background Targeted therapies for triple-negative breast cancer (TNBC) are limited; however, the epidermal growth factor receptor (EGFR) represents a potential target, as the majority of TNBC express EGFR. The purpose of these studies was to evaluate the effectiveness of two EGFR-targeted antibody-drug conjugates (ADC: ABT-414; ABBV-321) in combination with navitoclax, an antagonist of the anti-apoptotic BCL-2 and BCL-XL proteins, in order to assess the translational relevance of these combinations for TNBC. Methods The pre-clinical efficacy of combined treatments was evaluated in multiple patient-derived xenograft (PDX) models of TNBC. Microscopy-based dynamic BH3 profiling (DBP) was used to assess mitochondrial apoptotic signaling induced by navitoclax and/or ADC treatments, and the expression of EGFR and BCL-2/XL was analyzed in 46 triple-negative patient tumors. Results Treatment with navitoclax plus ABT-414 caused a significant reduction in tumor growth in five of seven PDXs and significant tumor regression in the highest EGFR-expressing PDX. Navitoclax plus ABBV-321, an EGFR-targeted ADC that displays more effective wild-type EGFR-targeting, elicited more significant tumor growth inhibition and regressions in the two highest EGFR-expressing models evaluated. The level of mitochondrial apoptotic signaling induced by single or combined drug treatments, as measured by DBP, correlated with the treatment responses observed in vivo. Lastly, the majority of triple-negative patient tumors were found to express EGFR and co-express BCL-XL and/or BCL-2. Conclusions The dramatic tumor regressions achieved using combined agents in pre-clinical TNBC models underscore the abilities of BCL-2/XL antagonists to enhance the effectiveness of EGFR-targeted ADCs and highlight the clinical potential for usage of such targeted ADCs to alleviate toxicities associated with combinations of BCL-2/XL inhibitors and systemic chemotherapies.

The molecular, histopathological, genomic and transcriptomic characteristics of uveal melanoma (UM) have identified four molecular subgroups with different clinical outcomes. Despite the improvements in UM classification and biological pathology, current treatments do not reduce the occurrence of metastasis. The development of effective adjuvant and metastatic therapies for UM has been slow and extremely limited. Preclinical models that closely resemble the molecular and genetic UM subgroups are essential for translating molecular findings into improved clinical treatment. In this review, we provide a retrospective view of the existing preclinical models used to study UM, and give an overview of their strengths and limitations. We review targeted therapy clinical trial data to evaluate the gap in the translation of preclinical findings to human studies. Reflecting on the current high attrition rates of clinical trials for UM, preclinical models that effectively recapitulate the human in vivo situation and/or accurately reflect the subtype classifications would enhance the translational impact of experimental data and have crucial implications for the advancement of personalised medicine.

Acute myeloid leukaemia (AML) is a heterogeneous haematopoietic malignancy caused by recurrent mutations in haematopoietic stem and progenitor cells that affect both the epigenetic regulatory machinery and signalling molecules. The t(8;21) or RUNX1‐RUNX1T1 translocation generates the RUNX1‐ETO chimeric transcription factor which primes haematopoietic stem cells for further oncogenic mutational events that in their sum cause overt disease. Significant progress has been made in generating both in vitro and in vivo model systems to recapitulate t(8;21) AML which are crucial for the understanding of the biology of the disease and the development of effective treatment. This review provides a comprehensive overview of the in vivo and in vitro model systems that were developed to gain insights into the molecular mechanisms of RUNX1‐ETO oncogenic activity and their contribution to the advancement of knowledge in the t(8;21) AML field. Such models include transgenic mice, patient‐derived xenografts, RUNX1‐ETO transduced human progenitor cells, cell lines and human embryonic stem cell model systems, making the t(8;21) as one of the well‐characterized sub‐type of AML at the molecular level., The t(8;21) or RUNX1‐RUNX1T1 translocation generates the RUNX1/ETO chimeric transcription factor which in cooperation with other mutations drives one of the most frequent types of acute myeloid leukaemia. Here, we provide a comprehensive overview of the in vivo and in vitro model systems that informed about the molecular mechanisms of RUNX1/ETO oncogenic activity.

Background Cancer is a highly heterogeneous disease with varying responses to anti-cancer drugs. Although several attempts have been made to predict the anti-cancer therapeutic responses, there remains a great need to develop highly accurate prediction models of response to the anti-cancer drugs for clinical applications toward a personalized medicine. Patient derived xenografts (PDXs) are preclinical cancer models in which the tissue or cells from a patient’s tumor are implanted into an immunodeficient or humanized mouse. In the present study, we develop a bioinformatics analysis pipeline to build a predictive gene expression model (GEM) for cancer patients’ drug responses based on gene expression and drug activity data from PDX models. Results Drug sensitivity biomarkers were identified by performing an association analysis between gene expression levels and post-treatment tumor volume changes in PDX models. We built a drug response prediction model (called PDXGEM) in a random-forest algorithm by using a subset of the drug sensitvity biomarkers with concordant co-expression patterns between the PDXs and pretreatment cancer patient tumors. We applied the PDXGEM to several cytotoxic chemotherapies as well as targeted therapy agents that are used to treat breast cancer, pancreatic cancer, colorectal cancer, or non-small cell lung cancer. Significantly accurate predictions of PDXGEM for pathological response or survival outcomes were observed in extensive independent validations on multiple cancer patient datasets obtained from retrospective observational studies and prospective clinical trials. Conclusion Our results demonstrated the strong potential of using molecular profiles and drug activity data of PDX tumors in developing a clinically translatable predictive cancer biomarkers for cancer patients. The PDXGEM web application is publicly available at http://pdxgem.moffitt.org.

New methods of tumor ablation have shown exciting efficacy in pre-clinical models but often demonstrate limited success in the clinic. Due to a lack of quality or quantity in primary malignant tissue specimens, therapeutic development and optimization studies are typically conducted on healthy tissue or cell-line derived rodent tumors that don't allow for high resolution modeling of mechanical, chemical, and biological properties. These surrogates do not accurately recapitulate many critical components of the tumor microenvironment that can impact in situ treatment success. Here, we propose utilizing patient-derived xenograft (PDX) models to propagate clinically relevant tumor specimens for the optimization and development of novel tumor ablation modalities. Specimens from three individual pancreatic ductal adenocarcinoma (PDAC) patients were utilized to generate PDX models. This process generated 15-18 tumors that were allowed to expand to 1.5 cm in diameter over the course of 50-70 days. The PDX tumors were morphologically and pathologically identical to primary tumor tissue. Likewise, the PDX tumors were also found to be physiologically superior to other in vitro and ex vivo models based on immortalized cell lines. We utilized the PDX tumors to refine and optimize irreversible electroporation (IRE) treatment parameters. IRE, a novel, non-thermal tumor ablation modality, is being evaluated in a diverse range of cancer clinical trials including pancreatic cancer. The PDX tumors were compared against either Pan02 mouse derived tumors or resected tissue from human PDAC patients. The PDX tumors demonstrated similar changes in electrical conductivity and Joule heating following IRE treatment. Computational modeling revealed a high similarity in the predicted ablation size of the PDX tumors that closely correlate with the data generated with the primary human pancreatic tumor tissue. Gene expression analysis revealed that IRE treatment resulted in an increase in biological pathway signaling associated with interferon gamma signaling, necrosis and mitochondria dysfunction, suggesting potential co-therapy targets. Together, these findings highlight the utility of the PDX system in tumor ablation modeling for IRE and increasing clinical application efficacy. It is also feasible that the use of PDX models will significantly benefit other ablation modality testing beyond IRE. Virginia-Maryland College of Veterinary Medicine; Virginia Tech Institute for Critical Technology and Applied Science Center for Engineered Health; National Institutes of HealthUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [R21EB028429]; AngioDynamics; American Association of Immunologist Careers in Immunology Fellowship Program This work was supported by the Virginia-Maryland College of Veterinary Medicine (IA), the Virginia Tech Institute for Critical Technology and Applied Science Center for Engineered Health (IA), National Institutes of Health R21EB028429 (IA), and AngioDynamics (IA, RD). Student work on this publication was supported by the American Association of Immunologist Careers in Immunology Fellowship Program (VR-S).

Background Chimeric antigen receptor T cells (CAR-T cells) therapy has been well recognized for treating B cell-derived malignancy. However, the efficacy of CAR-T cells against solid tumors remains dissatisfactory, partially due to the heterogeneity of solid tumors and T cell exhaustion in tumor microenvironment. PD-L1 is up-regulated in multiple solid tumors, resulting in T cell exhaustion upon binding to its receptor PD-1. Methods Here, we designed a dominant-negative form of PD-1, dPD1z, a vector containing the extracellular and transmembrane regions of human PD-1, and a CAR vector against PD-L1, CARPD-L1z, a vector employs a high-affinity single-chain variable fragment (scFv) against human PD-L1. These two vectors shared the same intracellular structure, including 4-1BB and TLR2 co-stimulatory domains, and the CD3ζ signaling domain. Results dPD1z T and CARPD-L1z T cells efficiently lysed PD-L1+ tumor cells and had enhanced cytokine secretion in vitro and suppressed the growth of non-small cell lung cancer (NSCLC), gastric cancer and hepatoma carcinoma in patient-derived xenograft (PDX). However, the combination of anti-mesothelin CAR-T cells (CARMSLNz T) with dPD1z T or CARPD-L1z T cells did not repress tumor growth synergistically in PDX, as CARMSLNz T cells upregulated PD-L1 expression upon activation and were subsequently attacked by dPD1z T or CARPD-L1z T cells. Conclusions In conclusion, we demonstrate CAR-T cells targeting PD-L1 were effective for suppressing the growth of multiple types of solid tumors in PDX models though their safety needs to be carefully examined.

BackgroundHundreds of systemic chemotherapy trials in diffuse intrinsic pontine glioma (DIPG) have not improved survival, potentially due to lack of intratumoral penetration, which has not previously been assessed in humans.MethodsWe used gemcitabine as a model agent to assess DIPG intratumoral pharmacokinetics (PK) using mass spectrometry.ResultsIn a phase 0 clinical trial of i.v. gemcitabine prior to biopsy in children newly diagnosed with DIPG by MRI, mean concentration in 4 biopsy cores in patient 1 (H3K27M diffuse midline glioma) was 7.65 µM. These compare favorably to levels for patient 2 (mean 3.85 µM, found to have an H3K27-wildtype low-grade glioma on histology), and from a similar study in adult glioblastoma (adjusted mean 3.48 µM). In orthotopic patient-derived xenograft (PDX) models of DIPG and H3K27M-wildtype pediatric glioblastoma, gemcitabine levels and clearance were similar in tumor, pons, and cortex and did not depend on H3K27 mutation status or tumor location. Normalized gemcitabine levels were similar in patient 1 and the DIPG PDX.ConclusionsThese findings, while limited to one agent, provide preliminary evidence for the hypotheses that lack of intratumoral penetration is not why systemic chemotherapy has failed in DIPG, and orthotopic PDX models can adequately model intratumoral PK in human DIPG.

[Purpose] Epigenetic regulation is crucial in mammalian development and maintenance of tissue-cell specific functions. Perturbation of epigenetic balance may lead to alterations in gene expression, resulting in cellular transformation and malignancy. Previous studies in Ewing sarcoma (ES) have shown that the Nucleosome Remodeling Deacetylase (NuRD) complex binds directly to EWS-FLI1 oncoprotein and modulates its transcriptional activity. The role of EWS-FLI1 as a driver of proliferation and transformation in ES is widely known, but the effect of epigenetic drugs on fusion activity remains poorly described. The present study evaluated the combination effects of the histone deacetylases inhibitor suberoylanilide hydroxamic acid (SAHA) and Lysine-specific demethylase1 inhibitor (HCI-2509) on different biological functions in ES and in comparison to monotherapy treatments., [Results] The study of proliferation and cell viability showed a synergistic effect in most ES cell lines analyzed. An enhanced effect was also observed in the induction of apoptosis, together with accumulation of cells in G1 phase and a blockage of the migratory capacity of ES cell lines. Treatment, either in monotherapy or in combination, caused a significant decrease of EWS-FLI1 mRNA and protein levels and this effect is mediated in part by fusion gene promoter regulation. The anti-tumor effect of this combination was confirmed in patient-derived xenograft mouse models, in which only the combination treatment led to a statistically significant decrease in tumor volume., [Conclusions] The combination of SAHA and HCI-2509 is proposed as a novel treatment strategy for ES patients to inhibit the essential driver of this sarcoma and tumor growth., Research in EDA, JM, and OMT labs is supported by Asociación Española Contra el Cáncer (AECC). Enrique de Alava’s lab is also supported by the Ministry of Economy and Competitiveness of Spain-FEDER (CIBERONC, PI11700464, RD06/0020/0059, FMGE) and the European Commission (FP7-HEALTH-2011-two-stage, Project ID278742 EUROSARC). DGD and LHP are supported by Red Temática de Investigación Cooperativa en Cáncer (RD12/0036/0017) and Centro de Investigación Biomédica en Red de Cáncer (CB16/12/00361)S. PRN is supported by a grant from the María García Estrada Foundation. DGD is a PhD researcher funded by the Consejería de Salud, Junta de Andalucía (PI-0197-2016). MJR is supported by the Consejería de Salud, Junta de Andalucía (ECAIF2-0176-2013).

Breast cancer is a complex disease in which each patient could present several genetic alterations that are therapeutically relevant in cancers. Here we explored the therapeutic benefit of combining PARP and mTOR inhibitors in a context of DNA repair deficiency and PI3K pathway activation. The combination of everolimus and olaparib was tested in BRCA2-mutated patient-derived xenografts (PDX) carrying alterations in the PI3K/AKT/mTOR pathway. An RPPA analysis of different signalling pathways was performed in untreated and treated xenografts. Everolimus and olaparib showed marked anti-tumor activities in the monotherapy setting and high efficacy when given in combination with 100% of mice showing tumor regressions. The fraction of P-H2AX positive cells was increased in both monotherapy arms and strongly increased in the combination setting. Everolimus given as monotherapy resulted in downregulation of different proteins involved in DNA damage repair, including FANCD2, RAD50 and SUV39H1. In the combination setting, expression of these proteins was almost completely abolished, suggesting convergence of PARP and mTOR in downregulation of DNA damage repair components. In conclusion, our results suggest that combining mTOR and DNA repair inhibition could be a successful strategy to treat a subset of breast cancer with BRCA2 mutation and alterations in the PI3K/AKT/mTOR pathway.

Wntless/GPR177 functions as WNT ligand carrier protein and activator of WNT/β-catenin signaling, however, its molecular role in gastric cancer (GC) has remained elusive. We investigated the role of GPR177 in gastric tumorigenesis and provided the therapeutic potential of a clinical development of anti-GPR177 monoclonal antibodies. GPR177 mRNA expression was assessed in GC transcriptome data sets (GSE15459, n = 184; GSE66229, n = 300); protein expression was assessed in independent patient tumor tissues (Yonsei TMA, n = 909). GPR177 expression were associated with unfavorable prognosis [log-rank test, GSE15459 (P = 0.00736), GSE66229 (P = 0.0142), and Yonsei TMA (P = 0.0334)] and identified as an independent risk predictor of clinical outcomes: GSE15459 [hazard ratio (HR) 1.731 (95% confidence interval; CI; 1.103- 2.715), P = 0.017], GSE66229 [HR 1.54 (95% CI, 1.10-2.151), P = 0.011], and Yonsei TMA [HR 1.254 (95% CI, 1.049- 1.500), P = 0.013]. Either antibody treatment or GPR177 knockdown suppressed proliferation of GC cells and sensitized cells to apoptosis. And also inhibition of GPR177 suppresses in vitro and in vivo tumorogenesis in GC cells and inhibits WNT/β-catenin signaling. Finally, targeting and inhibition of GPR177 with antibody suppressed tumorigenesis in PDX model. Together, these results suggest GPR177 as a novel candidate for prognostic marker as well as a promising target for treatment of GC patients. [BMB Reports 2018; 51(5): 255-260].

Heparanase, the sole heparan sulfate (HS) degrading endoglycosidase, regulates multiple biological activities that enhance tumor growth, metastasis, angiogenesis, and inflammation. Heparanase accomplishes this by degrading HS and thereby facilitating cell invasion and regulating the bioavailability of heparin-binding proteins. HS mimicking compounds that inhibit heparanase enzymatic activity were examined in numerous preclinical cancer models. While these studies utilized established tumor cell lines, the current study utilized, for the first time, patient-derived xenografts (PDX) which better resemble the behavior and drug responsiveness of a given cancer patient. We have previously shown that heparanase levels are substantially elevated in lung cancer, correlating with reduced patients survival. Applying patient-derived lung cancer xenografts and a potent inhibitor of heparanase enzymatic activity (PG545) we investigated the significance of heparanase in the pathogenesis of lung cancer. PG545 was highly effective in lung cancer PDX, inhibiting tumor growth in >85% of the cases. Importantly, we show that PG545 was highly effective in PDX that did not respond to conventional chemotherapy (cisplatin) and vice versa. Moreover, we show that spontaneous metastasis to lymph nodes is markedly inhibited by PG545 but not by cisplatin. These results reflect the variability among patients and strongly imply that PG545 can be applied for lung cancer therapy in a personalized manner where conventional chemotherapy fails, thus highlighting the potential benefits of developing anti-heparanase treatment modalities for oncology.

BACKGROUND: While it has been challenging to establish prostate cancer patient-derived xenografts (PDXs), with a take rate of 10-40% and long latency time, multiple groups throughout the world have developed methods for the successful establishment of serially transplantable human prostate cancer PDXs using a variety of immune deficient mice. In 2014, the Movember Foundation launched a Global Action Plan 1 (GAP1) project to support an international collaborative prostate cancer PDX program involving eleven groups. Between these Movember consortium members, a total of 98 authenticated human prostate cancer PDXs were available for characterization. Eighty three of these were derived directly from patient material, and 15 were derived as variants of patient-derived material via serial passage in androgen deprived hosts. A major goal of the Movember GAP1 PDX project was to provide the prostate cancer research community with a summary of both the basic characteristics of the 98 available authenticated serially transplantable human prostate cancer PDX models and the appropriate contact information for collaborations. Herein, we report a summary of these PDX models. METHODS: PDX models were established in immunocompromised mice via subcutaneous or subrenal-capsule implantation. Dual-label species (ie, human vs mouse) specific centromere and telomere Fluorescence In Situ Hybridization (FISH) and immuno-histochemical (IHC) staining of tissue microarrays (TMAs) containing replicates of the PDX models were used for characterization of expression of a number of phenotypic markers important for prostate cancer including AR (assessed by IHC and FISH), Ki67, vimentin, RB1, P-Akt, chromogranin A (CgA), p53, ERG, PTEN, PSMA, and epithelial cytokeratins. RESULTS: Within this series of PDX models, the full spectrum of clinical disease stages is represented, including androgen-sensitive and castration-resistant primary and metastatic prostate adenocarcinomas as well as prostate carcinomas with neuroendocrine differentiation. The annotated clinical characteristics of these PDXs were correlated with their marker expression profile. CONCLUSION: Our results demonstrate the clinical relevance of this series of PDXs as a platform for both basic science studies and therapeutic discovery/drug development. The present report provides the prostate cancer community with a summary of the basic characteristics and a contact information for collaborations using these models.

Background/Aims: Effective drug treatment for intrahepatic cholangiocarcinoma (ICC) is currently lacking. Therefore, there is an urgent need for new targets and new drugs that can prolong patient survival. Recently targeting the ubiquitin proteasome pathway has become an attractive anti-cancer strategy. In this study, we aimed to evaluate the therapeutic effect of and identify the potential mechanisms involved in targeting the proteasome subunit ADRM1 for ICC. Methods: The expression of ADRM1 and its prognostic value in ICC was analyzed using GEO and TCGA datasets, tumor tissues, and tumor tissue arrays. The effects of RA190 on the proliferation and survival of both established ICC cell lines and primary ICC cells were examined in vitro. Annexin V/propidium iodide staining, western blotting and immunohistochemical staining were performed. The in vivo anti-tumor effect of RA190 on ICC was validated in subcutaneous xenograft and patient-derived xenograft (PDX) models. Results: ADRM1 levels were significantly higher in ICC tissues than in normal bile duct tissues. ICC patients with high ADRM1 levels had worse overall survival (hazard ratio [HR] = 2.383, 95% confidence interval [CI] =1.357 to 4.188) and recurrence-free survival (HR = 1.710, 95% CI =1.045 to 2.796). ADRM1 knockdown significantly inhibited ICC growth in vitro and in vivo. The specific inhibitor RA190 targeting ADRM1 suppressed proliferation and reduced cell vitality of ICC cell lines and primary ICC cells significantly in vitro. Furthermore, RA190 significantly inhibited the proteasome by inactivating ADRM1, and the consequent accumulation of ADRM1 substrates decreased the activating levels of NF-κB to aggravate cell apoptosis. The therapeutic benefits of RA190 treatment were further demonstrated in both subcutaneous implantation and PDX models. Conclusions: Our findings indicate that up-regulated ADRM1 was involved in ICC progression and suggest the potential clinical application of ADRM1 inhibitors (e.g., RA190 and KDT-11) for ICC treatment.

Background: While patient-derived xenografts (PDXs) offer a powerful modality for translational cancer research, a precise evaluation of how accurately patient responses correlate with matching PDXs in a large, heterogeneous population is needed for assessing the utility of this platform for preclinical drug-testing and personalized patient cancer treatment. Patients and methods: Tumors obtained from surgical or biopsy procedures from 237 cancer patients with a variety of solid tumors were implanted into immunodeficient mice and whole-exome sequencing was carried out. For 92 patients, responses to anticancer therapies were compared with that of their corresponding PDX models. Results: We compared whole-exome sequencing of 237 PDX models with equivalent information in The Cancer Genome Atlas database, demonstrating that tumorgrafts faithfully conserve genetic patterns of the primary tumors. We next screened PDXs established for 92 patients with various solid cancers against the same 129 treatments that were administered clinically and correlated patient outcomes with the responses in corresponding models. Our analysis demonstrates that PDXs accurately replicate patients' clinical outcomes, even as patients undergo several additional cycles of therapy over time, indicating the capacity of these models to correctly guide an oncologist to treatments that are most likely to be of clinical benefit. Conclusions: Integration of PDX models as a preclinical platform for assessment of drug efficacy may allow a higher success-rate in critical end points of clinical benefit.

// Saqer F. Alsaqer 1,* , Mustafa M. Tashkandi 1,* , Vinay K. Kartha 2,3 , Ya-Ting Yang 1 , Yazeed Alkheriji 1 , Andrew Salama 4 , Xaralabos Varelas 5 , Maria Kukuruzinska 1 , Stefano Monti 2,3 and Manish V. Bais 1 1 Department of Molecular and Cell Biology, Boston University Henry M. Goldman School of Dental Medicine, Boston, MA, USA 2 Bioinformatics Program, Boston University, Boston, MA, USA 3 Section of Computational Biomedicine, Boston University School of Medicine, Boston, MA, USA 4 Department of Oral and Maxillofacial Surgery, Boston University Henry M. Goldman School of Dental Medicine, Boston, MA, USA 5 Department of Biochemistry, Boston University School of Medicine, Boston, MA, USA * These authors have contributed equally to this work Correspondence to: Manish V. Bais, email: // Keywords : LSD1, CTGF, OSCC, PDX, orthotopic mouse model Received : December 28, 2016 Accepted : July 14, 2017 Published : July 27, 2017 Abstract Lysine-specific demethylase 1 (LSD1) is a nuclear histone demethylase and a member of the amine oxidase (AO) family. LSD1 is a flavin-containing AO that specifically catalyzes the demethylation of mono- and di-methylated histone H3 lysine 4 through an FAD-dependent oxidative reaction. LSD1 is inappropriately upregulated in lung, liver, brain and esophageal cancers, where it promotes cancer initiation, progression, and metastasis. However, unlike other lysine-specific demethylases, the role and specific targets of LSD1 in oral squamous cell carcinoma (OSCC) pathogenesis remain unknown. We show that LSD1 protein expression was increased in malignant OSCC tissues in a clinical tissue microarray, and its expression correlated with progressive tumor stages. In an orthotopic oral cancer mouse model, LSD1 overexpression in aggressive HSC-3 cells promoted metastasis whereas knockdown of LSD1 inhibited tumor spread, suggesting that LSD1 is a key regulator of OSCC metastasis. Pharmacological inhibition of LSD1 using a specific small molecule inhibitor, GSK-LSD1, down-regulated EGF signaling pathway. Further, GSK-LSD1 attenuates CTGF/CCN2, MMP13, LOXL4 and vimentin expression but increased E-cadherin expression in pre-existing, patient-derived tonsillar OSCC xenografts. Similarly, GSK-LSD1 inhibited proliferation and CTGF expression in mesenchymal cells, including myoepithelial cells and osteosarcoma cells. In addition, gene set enrichment analysis revealed that GSK-LSD1 increased p53 expression and apoptosis while inhibiting c-myc, β-catenin and YAP-induced oncogenic transcriptional networks. These data reveal that aberrant LSD1 activation regulates key OSCC microenvironment and EMT promoting factors, including CTGF, LOXL4 and MMP13.

Research using animal model systems has been instrumental in delivering improved therapies for breast cancer, as well as in generating new insights into the mechanisms that underpin development of the disease. A large number of different models are now available, reflecting different types and stages of the disease; choosing which one to use depends on the specific research question(s) to be investigated. Based on presentations and discussions from leading experts who attended a recent workshop focused on in vivo models of breast cancer, this article provides a perspective on the many varied uses of these models in breast cancer research, their strengths, associated challenges and future directions. Among the questions discussed were: how well do models represent the different stages of human disease; how can we model the involvement of the human immune system and microenvironment in breast cancer; what are the appropriate models of metastatic disease; can we use models to carry out preclinical drug trials and identify pathways responsible for drug resistance; and what are the limitations of patient-derived xenograft models? We briefly outline the areas where the existing breast cancer models require improvement in light of the increased understanding of the disease process, reflecting the drive towards more personalised therapies and identification of mechanisms of drug resistance., Summary: This Review provides a summary of the many varied uses of mouse models in breast cancer research focusing on their strengths, challenges and future directions.

Targeted therapies in endometrial cancer (EC) using kinase inhibitors rarely result in complete tumor remission and are frequently challenged by the appearance of refractory cell clones, eventually resulting in disease relapse. Dissecting adaptive mechanisms is of vital importance to circumvent clinical drug resistance and improve the efficacy of targeted agents in EC. Sorafenib is an FDA-approved multitarget tyrosine and serine/threonine kinase inhibitor currently used to treat hepatocellular carcinoma, advanced renal carcinoma and radioactive iodine-resistant thyroid carcinoma. Unfortunately, sorafenib showed very modest effects in a multi-institutional phase II trial in advanced uterine carcinoma patients. Here, by leveraging RNA-sequencing data from the Cancer Cell Line Encyclopedia and cell survival studies from compound-based high-throughput screenings we have identified the lysosomal pathway as a potential compartment involved in the resistance to sorafenib. By performing additional functional biology studies we have demonstrated that this resistance could be related to macroautophagy/autophagy. Specifically, our results indicate that sorafenib triggers a mechanistic MAPK/JNK-dependent early protective autophagic response in EC cells, providing an adaptive response to therapeutic stress. By generating in vivo subcutaneous EC cell line tumors, lung metastatic assays and primary EC orthoxenografts experiments, we demonstrate that targeting autophagy enhances sorafenib cytotoxicity and suppresses tumor growth and pulmonary metastasis progression. In conclusion, sorafenib induces the activation of a protective autophagic response in EC cells. These results provide insights into the unopposed resistance of advanced EC to sorafenib and highlight a new strategy for therapeutic intervention in recurrent EC. This work was supported by MINECO (SAF2016-80157-R), the Fondo de Investigaciones Sanitarias (PI10/00922, PI10/00604, PI13/01339 and PIE13–00022 (Oncoprofile)), Grant 2009SGR794 (Barcelona, Spain), Fundaci on Asociaci on Espa~nola Contra el C ancer (AECC-2011), AECC_- Barcelona and RETICS (RD12/0036/0013). DLN is recipient of a NURF scheme. The funders had no involvement in the study design, execution, analysis or interpretation of data and writing of the manuscript.

Background: Screening of patients for cancer-driving mutations is now used for cancer prognosis, remission scoring and treatment selection. Although recently emerged targeted next-generation sequencing-based approaches offer promising diagnostic capabilities, there are still limitations. There is a pressing clinical need for a well-validated, rapid, cost-effective mutation profiling system in patient specimens. Given their speed and cost-effectiveness, quantitative PCR mutation detection techniques are well suited for the clinical environment. The qBiomarker mutation PCR array has high sensitivity and shorter turnaround times compared with other methods. However, a direct comparison with existing viable alternatives are required to assess its true potential and limitations. Methods: In this study, we evaluated a panel of 117 patient-derived tumour xenografts by the qBiomarker array and compared with other methods for mutation detection, including Ion AmpliSeq sequencing, whole-exome sequencing and droplet digital PCR. Results: Our broad analysis demonstrates that the qBiomarker's performance is on par with that of other labour-intensive and expensive methods of cancer mutation detection of frequently altered cancer-associated genes, and provides a foundation for supporting its consideration as an option for molecular diagnostics. Conclusions: This large-scale direct comparison and validation of currently available mutation detection approaches is extremely relevant for the current scenario of precision medicine and will lead to informed choice of screening methodologies, especially in lower budget conditions or time frame limitations.

Preclinical imaging is critical in the development of translational strategies to detect diseases and monitor response to therapy. The National Cancer Institute Co-Clinical Imaging Resource Program was launched, in part, to develop best practices in preclinical imaging. In this context, the objective of this work was to develop a 1-hour, multiparametric magnetic resonance image-acquisition pipeline with triple-negative breast cancer patient-derived xenografts (PDXs). The 1-hour, image-acquisition pipeline includes T1- and T2-weighted scans, quantitative T1, T2, and apparent diffusion coefficient (ADC) parameter maps, and dynamic contrast-enhanced (DCE) time-course images. Quality-control measures used phantoms. The triple-negative breast cancer PDXs used for this study averaged 174 ± 73 μL in volume, with region of interest–averaged T1, T2, and ADC values of 1.9 ± 0.2 seconds, 62 ± 3 milliseconds, and 0.71 ± 0.06 μm2/ms (mean ± SD), respectively. Specific focus was on assessing the within-subject test–retest coefficient-of-variation (CVWS) for each of the magnetic resonance imaging metrics. Determination of PDX volume via manually drawn regions of interest is highly robust, with ∼1% CVWS. Determination of T2 is also robust with a ∼3% CVWS. Measurements of T1 and ADC are less robust with CVWS values in the 6%–11% range. Preliminary DCE test–retest time-course determinations, as quantified by area under the curve and Ktrans from 2-compartment exchange (extended Tofts) modeling, suggest that DCE is the least robust protocol, with ∼30%–40% CVWS.

Patient-Derived Xenografts (PDXs) are, so far, the best preclinical model to validate targets and predictors of response to therapy. While subcutaneous implantation very rarely allows metastatic dissemination, orthotopic implantation (Patient-Derived Orthotopic Xenograft-PDOX) increases metastatic capability. Using a modified tool to analyze model validity, we performed a systematic review of Embase, PubMed, and Web of Science up to December 2018 to identify all original publications describing gastric cancer (GC) PDOXs. We identified ten studies of PDOX model validation from January 1981 to December 2018 that fulfilled the inclusion and exclusion criteria. Most models (70%) were derived from human GC cell lines rather than tissue fragments. In 90% of studies, the implantation was performed in the subserosal layer. Tumour engraftment rate ranged from 0 to 100%, despite the technique. Metastases were observed in 40% of PDOX models implanted into the subserosal layer, employing either cell suspension or cell line-derived tumour fragments. According to our modified model validity tool, half of the studies were defined as unclear because one or more validation criteria were not reported. Available GC PDOX models are not adequate according to our model validity tool. There is no demonstration that the submucosal site is more effective than the subserosal layer, and that tissue fragments are better than cell suspensions for successful engraftment and metastatic spread. Further studies should strictly employ model validity tools and large samples with orthotopic implant sites mirroring as much as possible the donor tumour characteristics.

Glioblastoma (GBM) is the most aggressive primary brain tumor in adults. Designing effective individualized therapies for GBM requires quality fresh tissue specimens, and a comprehensive molecular profile of this highly heterogenous neoplasm. Novel neuro-surgical approaches, such as the automated resection NICO Myriad™ system, are increasingly used by neurosurgeons to better reach the invasive front of tumors. However, no information exists on how harvesting GBM tissue using this approach may impact the translational research value of the sample. Here, we set out to characterize matched specimens from 15 patients, where one tissue sample was obtained using traditional tumor de-bulking (herein referred to as “en bloc” sample), and the other sample was obtained using the MyriadTM System (herein referred to as “Myriad” sample). We investigated the fidelity of patient derived xenografts (PDXs) for each sample type to the corresponding human tissues and evaluated the added value of sequencing both samples for each patient. Matched en bloc and Myriad samples processed in parallel, were subjected to the following assays: cell viability, self-renewal, in vivo tumorigenicity using an orthotopic model of glioma, genomic sequencing, and pharmacological testing using PI3K-MTOR pathway inhibitors. Our results demonstrate that primary GBM cultures derived from matched specimens grew at similar rates (correlation coefficient R = 0.72), generated equivalent number of neurospheres, and had equivalent tumorigenic potential in vivo (mouse survival correlation coefficient R = 0.93). DNA Sequencing using the Illumina tumor panel amplicons revealed over 70% concordance in non-synonymous mutations between matched human GBM specimens. PDX genomic profiles were also highly concordant with the corresponding patient tissues (>70%). RNA sequencing of paired GBM samples revealed unique genomic variants and differential gene expression between the en bloc and Myriad specimens, with the former molecularly resembling the “tumor core” and the latter resembling the “invasive tumor front” signature. Functionally, we show that primary-derived GBM cells—obtained after fresh specimen's dissociation—are more effectively growth-inhibited by co-targeting non-overlapping mutations enriched in each sample type, suggesting that profiling both specimens more adequately capture the molecular heterogeneity of GBM and may enhance the design accuracy and efficacy of individualized therapies.

In up to 30% of non-small cell lung cancer (NSCLC) patients, the oncogenic driver of tumor growth is a constitutively activated epidermal growth factor receptor (EGFR). Although these patients gain great benefit from treatment with EGFR tyrosine kinase inhibitors, the development of resistance is inevitable. To model the emergence of drug resistance, an EGFR-driven, patient-derived xenograft (PDX) NSCLC model was treated continuously with Gefitinib in vivo. Over a period of more than three months, three separate clones developed and were subsequently analyzed: Whole exome sequencing and reverse phase protein arrays (RPPAs) were performed to identify the mechanism of resistance. In total, 13 genes were identified, which were mutated in all three resistant lines. Amongst them the mutations in NOMO2, ARHGEF5 and SMTNL2 were predicted as deleterious. The 53 mutated genes specific for at least two of the resistant lines were mainly involved in cell cycle activities or the Fanconi anemia pathway. On a protein level, total EGFR, total Axl, phospho-NF&kappa, B, and phospho-Stat1 were upregulated. Stat1, Stat3, MEK1/2, and NF&kappa, B displayed enhanced activation in the resistant clones determined by the phosphorylated vs. total protein ratio. In summary, we developed an NSCLC PDX line modelling possible escape mechanism under EGFR treatment. We identified three genes that have not been described before to be involved in an acquired EGFR resistance. Further functional studies are needed to decipher the underlying pathway regulation.

Human breast cancer is characterized by a high degree of inter-patients heterogeneity in terms of histology, genomic alterations, gene expression patterns, and metastatic behavior, which deeply influences individual prognosis and treatment response. The main cause of mortality in breast cancer is the therapy-resistant metastatic disease, which sets the priority for novel treatment strategies for these patients. In the present study, we demonstrate that Patient Derived Xenografts (PDXs) that were obtained from metastatic and therapy-resistant breast cancer samples recapitulate the wide spectrum of the disease in terms of histologic subtypes and mutational profiles, as evaluated by whole exome sequencing. We have integrated genomic and transcriptomic data to identify oncogenic and actionable pathways in each PDX. By taking advantage of primary short-term in vitro cultures from PDX tumors, we showed their resistance to standard chemotherapy (Paclitaxel), as seen in the patients. Moreover, we selected targeting drugs and analyzed PDX sensitivity to single agents or to combination of targeted and standard therapy on the basis of PDX-specific genomic or transcriptomic alterations. Our data demonstrate that PDXs represent a suitable model to test new targeting drugs or drug combinations and to prioritize personalized therapeutic regimens for pre-clinal and clinical tests.

Novel inhibitors are urgently needed for use as targeted therapies to improve the overall survival (OS) of patients with ovarian cancer. Here, we show that cell division cycle 25B (CDC25B) is over-expressed in ovarian tumors and associated with poor patient prognosis. All previously reported CDC25B inhibitors have been identified by their ability to reversibly inhibit the catalytic dephosphorylation activity of CDC25B in vitro; however, none of these compounds have entered clinical trials for ovarian cancer therapy. In this study, we synthesized a novel small molecule compound, WG-391D, that potently down-regulates CDC25B expression without affecting its catalytic dephosphorylation activity. The inhibition of CDC25B by WG-391D is irreversible, and WG-391D should therefore exhibit potent antitumor activity against ovarian cancer. WG-391D induces cell cycle progression arrest at the G2/M phase. Half maximal inhibitory concentration (IC50) values of WG-391D for inhibition of the proliferation and migration of eight representative ovarian cancer cell lines (SKOV3, ES2, OVCAR8, OVTOKO, A2780, IGROV1, HO8910PM, and MCAS) and five primary ovarian tumor cell lines (GFY004, GFY005, CZ001, CZ006, and CZ008) were lower than 10 and 1 μM, respectively. WG-391D inhibited tumor growth in nude mice inoculated with SKOV3 cells or a patient-derived xenograft (PDX). The underlying mechanisms were associated with the down-regulation of CDC25B and subsequent inactivation of cell division cycle 2 (CDC2) and the serine/threonine kinase, AKT. In conclusion, this study demonstrates that WG-391D exhibits strong antitumor activity against ovarian cancer and indicates that the down-regulation of CDC25B by inhibitors could provide a rationale for ovarian cancer therapy.

Background Programmed death-ligand 1 (PD-L1) expression in metastatic renal cell carcinoma (RCC) correlates with a worse prognosis, but whether it also predicts responsiveness to anti-PD-1/PD-L1 therapy remains unclear. Most studies of PD-L1 are limited by evaluation in primary rather than metastatic sites, and in biopsy samples, which may not be representative. These limitations may be overcome with immuno–positron emission tomography (iPET), an emerging tool allowing the detection of cell surface proteins with radiolabeled antibodies. Here, we report iPET studies of PD-L1 in a preclinical tumorgraft model of clear cell RCC (ccRCC) from a patient who had a favorable response to anti-PD-1 therapy. Case presentation A 49-year-old man underwent a cytoreductive nephrectomy in 2017 of a right kidney tumor invading into the adrenal gland that was metastatic to the lungs and a rib. Histological analyses revealed a ccRCC of ISUP grade 4 with extensive sarcomatoid features. IMDC risk group was poor. Within two hours of surgery, a tumor sample was implanted orthotopically into NOD/SCID mice. Consistent with an aggressive tumor, a renal mass was detected 18 days post-implantation. Histologically, the tumorgraft showed sarcomatoid differentiation and high levels of PD-L1, similar to the patient’s tumor. PD-L1 was evaluated in subsequently transplanted mice using iPET and the results were compared to control mice implanted with a PD-L1-negative tumor. We labeled atezolizumab, an anti-PD-L1 antibody with a mutant Fc, with zirconium-89. iPET revealed significantly higher 89Zr-atezolizumab uptake in index than control tumorgrafts. The patient was treated with high-dose IL2 initially, and subsequently with pazopanib, with rapidly progressive disease, but had a durable response with nivolumab. Conclusions To our knowledge, this is the first report of non-invasive detection of PD-L1 in renal cancer using molecular imaging. This study supports clinical evaluation of iPET to identify RCC patients with tumors deploying the PD-L1 checkpoint pathway who may be most likely to benefit from PD-1/PD-L1 disrupting drugs.

Summary Robust patient-derived platforms that recapitulate the cellular and molecular fingerprints of glioblastoma are crucial for developing effective therapies. Here, we describe a chemically defined protocol for 3D culture and propagation of glioblastoma in 3D gliospheres, patient-derived organoids (PDOs), mouse brain orthotopic xenografts (PDOXs), and downstream drug and immunofluorescence assays. This simple-to-follow protocol allows assessing drug sensitivity, on-target activity, and combined drug synergy. Promising therapies can then be validated in PDOXs for translation in precision medicine oncology trials. For complete details on the use and execution of this protocol, please refer to Chadwick et al. (2020) and Patrizii et al. (2018)., Graphical Abstract, Highlights • Reproducible generation of 3D cultured glioblastoma patient-derived organoids (PDOs) • Gliospheres and PDOs can be propagated and cryopreserved for downstream applications • Engrafted cells are imaged with MRI/BLI in mouse brain orthotopic xenografts • PDOs are used to assess synergy between drug combinations for combined targeted therapy, Robust patient-derived platforms that recapitulate the cellular and molecular fingerprints of glioblastoma are crucial for developing effective therapies. Here, we describe a chemically defined protocol for 3D culture and propagation of glioblastoma in 3D gliospheres, patient-derived organoids (PDOs), mouse brain orthotopic xenografts (PDOXs), and downstream drug and immunofluorescence assays. This simple-to-follow protocol allows assessing drug sensitivity, on-target activity, and combined drug synergy. Promising therapies can then be validated in PDOXs for translation in precision medicine oncology trials.