Patient-derived Xenograft Research Articles

Angioimmunoblastic T-cell lymphoma: Characterization of clonal T and B cells and a patient-derived xenograft study of coexisting T- and B-cell proliferation.

Angioimmunoblastic T-cell lymphoma (AITL) is a rare and aggressive lymphoma with a poor prognosis. AITL is associated with Epstein-Barr virus (EBV)-positive B cells in most cases, suggesting a possible role for the virus in the pathobiology of AITL. Cell lines from AITL patients do not exist and models of human AITL are needed. We aim to establish such a model and use it for preclinical therapeutic evaluation. Primary lymph node tissue from an AITL patient was used for tumor cell isolation and injection to NSG mice. The established patient-derived xenograft (PDX) model was characterized by immunophenotyping, whole-exome sequencing (WES), and T/B-cell receptor gene rearrangement studies. In vivo AITL PDX trials were performed with elotuzumab, romidepsin, and rituximab. An AITL PDX mouse model that includes a coexisting EBV+ B-cell proliferation was established. We confirmed clonal identity of the engrafted T cells with the primary T-lymphoma cells. WES on DNA from xenografted sorted T and B cells identified eight and three mutations previously reported in the COSMIC database, respectively. Primary tumor cells could be passaged serially in NSG mice with an increasing percentage of monoclonal B cells that mimic the human condition in which the clonal B-cell component in some cases may mask an underling T-cell lymphoma. In this PDX mouse study, single agent elotuzumab or rituximab significantly improved mice survival. Survival was further improved when elotuzumab or romidepsin was combined with rituximab. To our knowledge, this is the first molecular characterization of AITL model coexisting with associated EBV+ B cells, and use of such a PDX model for therapeutic evaluation of agents targeting both malignant T cells and B cells simultaneously.

Acquired vulnerability against EGF receptor inhibition in gastric cancer promoted by class I histone deacetylase inhibitor entinostat.

Histone deacetylase inhibitors (HDACi) have shown promising preclinical activity in gastric cancer cells; unfortunately, however, these could not be confirmed in clinical trials. This highlights the need for the identification of underlying reasons, which may also provide the basis for possible combination therapies. Here, we delineated the effects of HDACi on components of EGFR signalling in gastric cancer cells. We investigated entinostat effects on EGFR and amphiregulin (AREG) expression in various cell line- and primary patient tumor-based in vitro, ex vivo and in vivo models, on the mRNA and protein level. Based on these results, a combined entinostat plus EGFR inhibitor erlotinib treatment in vitro and in vivo was studied. Proteomics analyses in gastric cancer cells treated with entinostat revealed a marked upregulation of EGFR in the majority of cell lines and an even more robust induction of the EGFR ligand AREG. This was confirmed in a panel of different cell lines in vitro, in tumor tissue-slice cultures ex vivo and in cell line- or patient-derived tumor xenografts in mice. Since previous studies in other tumor entities showed a downregulation of EGFR by HDACi, our findings thus indicate essential differences in the adaptive response of gastric carcinoma cells. Moreover, our results provided the basis for combined entinostat + EGFR inhibitor (erlotinib) treatment, and indeed we demonstrate synergistic effects in combination therapy studies. Our findings establish the profound upregulation of the EGFR/AREG axis by entinostat as starting point for a rational combination therapy in gastric carcinoma.

Patient-derived xenograft model in cancer: establishment and applications.

The patient-derived xenograft (PDX) model is a crucial in vivo model extensively employed in cancer research that has been shown to maintain the genomic characteristics and pathological structure of patients across various subtypes, metastatic, and diverse treatment histories. Various treatment strategies utilized in PDX models can offer valuable insights into the mechanisms of tumor progression, drug resistance, and the development of novel therapies. This review provides a comprehensive overview of the establishment and applications of PDX models. We present an overview of the history and current status of PDX models, elucidate the diverse construction methodologies employed for different tumors, and conduct a comparative analysis to highlight the distinct advantages and limitations of this model in relation to other in vivo models. The applications are elucidated in the domain of comprehending the mechanisms underlying tumor development and cancer therapy, which highlights broad applications in the fields of chemotherapy, targeted therapy, delivery systems, combination therapy, antibody-drug conjugates and radiotherapy. Furthermore, the combination of the PDX model with multiomics and single-cell analyses for cancer research has also been emphasized. The application of the PDX model in clinical treatment and personalized medicine is additionally emphasized.

H19 promotes polarization and alternative splicing in tumor-associated macrophages, facilitating pancreatic cancer progression.

Tumor-associated macrophages (TAMs) play a crucial physiological role in the pancreatic tumor microenvironment. However, the role of long non-coding RNAs (lncRNAs) in TAMs within pancreatic tumors remains unclear. By lncRNA sequencing between TAMs and resident macrophages from normal tissues in pancreatic cancer, it is found that H19 is highly expressed in TAMs and is correlated with the prognosis and stages of pancreatic cancer. Constructing a co-culture model of THP-1 derived TAMs and pancreatic cancer cells, H19 promotes the polarization of TAMs towards the M2 phenotype and the secretion of IL-6, IL-10, and TGF-β, both in vivo and in vitro, indirectly enhancing pancreatic cancer proliferation and metastasis. Mechanistically, H19 competitively binds to the mRNA of YTHDC1 with MiR-107, and also interacts with the YTHDC1 protein, regulating the stability of SRSF1 and thereby affecting the alternative splicing of IL-6 and IL-10. Utilizing organoids and the patient-derived xenograft (PDX) model, it is found that ruxolitinib may represent a promising treatment option for PDAC patients with high H19 expression.

Inhibiting de novo lipogenesis identifies a therapeutic vulnerability in therapy-resistant colorectal cancer.

A significant clinical challenge in patients with colorectal cancer (CRC), which adversely impacts patient survival, is the development of therapy resistance leading to a relapse. Therapy resistance and relapse in CRC is associated with the formation of lipid droplets (LD) by stimulating de novo lipogenesis (DNL). However, the molecular mechanisms underlying the increase in DNL and the susceptibility to DNL-targeted therapies remain unclear. Our study demonstrates that colorectal drug-tolerant persister cells (DTPs) over-express Lipin1 (LPIN1), which facilitates the sequestration of free fatty acids into LDs. The increased expression is mediated by the ETS1-PTPN1-c-Src-CEBPβ pathway. Blocking the conversion of free fatty acids into LDs by treatment with statins or inhibiting lipin1 expression disrupts lipid homeostasis, leading to lipotoxicity and ferroptotic cell death in both DTPs and patient-derived organoids (PDOs) in vitro. Ferroptosis inhibitors or N-acetylcysteine (NAC) can alleviate lipid ROS and cell death resulting from lipin1 inhibition. This strategy also significantly reduces tumor growth in CRC DTP mouse xenograft and patient-derived xenograft (PDX) models. Our findings highlight a new metabolic vulnerability in CRC DTPs, PDO, and PDX models and provide a framework for the rational repurposing of statins. Targeting the phosphatidic acid (PA) to diacylglycerol (DAG) conversion to prevent lipid droplet formation could be an effective therapeutic approach for therapy-resistant CRC.

Phase 1/2 study of FOG-001, a first-in-class direct β-catenin:TCF inhibitor, in patients with colorectal cancer, hepatocellular carcinoma, and other locally advanced or metastatic solid tumors.

TPS322 Background: Wnt/B-catenin pathway activation, which is frequently present in most colorectal cancers and a variety of other solid tumors, is associated with poor prognosis and resistance to immunotherapy. FOG-001 is a Helicon peptide that competitively inhibits the interaction between β-catenin and the T-cell factor (TCF) family of transcription factors. Experiments in patient-derived-xenograft (PDX) models of colorectal cancer (CRC) and hepatocellular carcinoma (HCC) have demonstrated that FOG-001 can inhibit tumor growth and promote tumor regression when administered as monotherapy or in combination with immune checkpoint inhibitors or standard of care therapies, including bevacizumab and 5-FU. Methods: This first-in-human, Phase 1/2, multicenter, open-label, dose escalation (Part 1) and dose-expansion (Part 2) study evaluates the safety/tolerability, pharmacokinetics (PK), pharmacodynamics (PD), and anti-tumor effects of FOG-001 as monotherapy and in combination with other anti-cancer therapies. Eligible patients must have received at least one prior systemic anti-cancer therapy and either progressed on, not responded to, or be unfit for available therapies. In Part 1, FOG-001 is administered intravenously, either weekly or every other week, at escalating dose levels evaluated sequentially in a standard 3+3 design as monotherapy in patients with: (1a) microsatellite stable (MSS) CRC or any solid tumor with a documented Wnt/b-catenin pathway activating mutation (WPAM) and (1c) HCC with WPAM. Part 1b will evaluate PD effects in cohorts of approximately six patients with MSS CRC. Upon selection of the preliminary recommended Phase 2 dose from Part 1a for weekly dosing, combination dose escalation (1d) evaluates the safety/tolerability, PK and anti-tumor effects of FOG-001 in CRC patients combined with FOLFOX+bevacizumab, anti-PD-1/PD-L1 (also includes other solid tumors with known WPAM), or trifluridine/tipiracil+bevacizumab. Part 2 dose expansion evaluates FOG-001 monotherapy in MSS CRC, HCC, and other solid tumors with documented WPAM. Combination dose expansion evaluates the combinations initially evaluated in Part 1d. Primary endpoints are safety/tolerability and preliminary anti-tumor effects (objective response rate). Secondary endpoints are PK, PD, and other anti-tumor responses (e.g., best objective response, duration of response, and progression free survival). Enrollment in Parts 1 and 2 is ongoing in the USA. Clinical trial information: NCT05919264 .

Avutometinib plus cetuximab after chemotherapy in patients with KRAS -mutated metastatic colorectal cancer: Results of phase 1 dose escalation.

199 Background: Anti-EGFR antibodies like cetuximab are ineffective in KRAS mutated (mt) colorectal cancer (CRC) due to constitutive activation of downstream pathways. Avutometinib is a first in class, oral, novel, dual RAF/MEK inhibitor. In patient-derived xenograft models of KRAS mt CRC, the combination of an anti-EGFR antibody with avutometinib conferred stronger tumor growth inhibition than either agent alone. We present results from the dose escalation phase (1b) of an ongoing phase 1b/2 trial evaluating the combination avutometinib plus cetuximab in KRAS mt metastatic (m) CRC. Methods: This is a single-center, open-label, phase 1b/2 study evaluating the safety and combined tolerability of avutometinib and cetuximab. Patients received avutometinib 2.4mg (at DL0) or 3.2mg (at DL1) orally twice a week for 3 weeks out of a 4-week cycle + cetuximab 500mg/m 2 intravenously q2week. Patients with disease progression on, or intolerance to 5-FU, oxaliplatin, and irinotecan, and measurable disease per RECIST v1.1 were enrolled. The primary endpoint for phase 1b was to establish the maximum tolerated dose (MTD) and hence determine the recommended phase 2 dose (RP2D). Dose limiting toxicity (DLT) period was 28 days. A 3+3 dose escalation design was used with dose level (DLs) 0 (2.4mg avutometinib) as the starting level and one dose escalation to DL+1 (3.2mg avutometinib). Results: We report safety results of phase 1b. 10 patients, 7 at DL0 and 3 at DL+1, were enrolled. Median age was 52.5 yrs (range 41 – 75). Any grade (G) treatment related adverse events (TRAEs) were seen in all 10 patients. Most common TRAEs were acneiform rash (100%), chills (30%), diarrhea (30%), and fatigue (30%). G3 TRAEs at DL0 were seen in 4/7 (57.1%) patients and at DL+1 in 2/3 (66.7%) patients. No G4 TRAEs were seen. G3 TRAEs seen at DL0 are noted in the table. One patient from the initial three enrolled was not evaluable due to underdosing. Another patient was later found to not meet eligibility criteria, but they did clear the DLT period. Hence, 6/7 patients at DL0 were evaluated for AE assessment, of which 1 had a DLT. 3 additional patients were enrolled in DL+1. 2/3 patients in DL+1 had grade 3 toxicities (both DLTs) of rash. Hence DL0 was established as the MTD/RP2D. Conclusions: This is the first trial evaluating the combination avutometinib plus cetuximab in KRAS mt mCRC. RP2D for avutometinib was established at 2.4mg orally twice a week for 3 weeks out of a 4-week cycle when used in combination with q2week 500mg/m 2 cetuximab for KRAS mt mCRC. Enrollment continues currently for the dose expansion phase for efficacy assessment. Clinical trial information: NCT05200442 . G3 TRAEs at DL0 attributed to both drugs unless specified. G3 TRAE # of patients Acneiform rash 3/7 (42.9%) Hand-Foot syndrome 1/7 (14.3%) Extremity edema 1/7 (14.3%) Cellulitis 1/7 (14.3%) Sepsis 1/7 (14.3%) Anemia 1/7 (14.3%) Creatine phosphokinase elevation 1/7 (14.3% – avutometinib only)

Development of a novel molecular probe for visualizing mesothelin on the tumor via positron emission tomography.

Mesothelin (MSLN) is an antigen that is overexpressed in various cancers, and its interaction with tumor-associated cancer antigen 125 plays a multifaceted role in tumor metastasis. The serum MSLN expression level can be detected using enzyme-linked immunosorbent assay; however, non-invasive visualization of its expression at the tumor site is currently lacking. Therefore, the aim of this study was to develop a molecular probe for imaging MSLN expression through positron emission tomography (PET). VHH 269-H4 was obtained via immunization of llama using a fragment of MSLN from residue 360 to residue 597. S-2-(4-isothiocyanatobenzyl)-1,4,7-triazacyclononane-1,4,7-triacetic acid (p-SCN-Bn-NOTA) was conjugated to VHH 269-H4 to yield precursor NOTA 269-H4 for radiolabeling. The chelator-to-VHH ratio was determined by mass spectrometry. The binding kinetics of VHH 269-H4 and NOTA 269-H4 were measured by surface plasmon resonance. Flow cytometry was carried out using the anti-mesothelin monoclonal antibody Anetumab to select MSLN-positive and MSLN-negative cell lines. After radiolabeling, the radiochemical purity and in vitro stability were tested by radio-thin-layer chromatography and size exclusion chromatography, respectively. A saturation binding assay was conducted to measure the dissociation constant (Kd) of [68Ga]Ga-NOTA-269-H4. By mircoPET/CT imaging and biodistribution studies, the in vivo performances of the novel tracer were investigated in NCG mice bearing OVCAR-8, SKOV-3, or patient-derived xenografts. VHH 269-H4 targeting MSLN was obtained with a Kd value of 0.3 nM. After conjugation, approximately 27% and 3.2% of VHH were coupled to one and two NOTA chelators, respectively. This yielded precursor NOTA 269-H4 with a Kd value of 1.1 nM. The radiochemistry was accomplished with moderate radiochemical yields (34 ± 14%, n = 9, decay-corrected). [68Ga]Ga-NOTA-269-H4 was obtained with high radiochemical purity (> 99%), and was stable after 90min incubation at room temperature. The binding affinity of the radioligand towards MSLN was kept in the nanomolar range. Flow cytometry revealed that OVCAR-8 cells possess a high level of MSLN expression, while MSLN expression on SKOV-3 cells was negligible. Consistently, in microPET/CT imaging, [68Ga]Ga-NOTA-269-H4 demonstrated clear tumor visualization using NCG mice bearing OVCAR-8 xenografts, but no radioactivity accumulation was observed in SKOV-3 xenografts, suggesting a high specificity of the tracer in vivo. In biodistribution studies, [68Ga]Ga-NOTA-269-H4 displayed radioactivity accumulation of 2.93 ± 0.39%ID/g in OVCAR-8 xenografts at 30min post-injection, and the highest tumor-to-blood ratio (~ 3) was achieved at 90min post-injection. In NCG mice bearing patient-derived xenografts, [68Ga]Ga-NOTA-269-H4 was able to noninvasively detect MSLN expression via microPET/CT imaging. To our knowledge, our studies achieved the first-time to non-invasively detect MSLN expression clearly using a single domain antibody fragment. To sum up, [68Ga]Ga-NOTA-269-H4 is a highly promising PET probe to visualize MSLN expression in vivo and holds great potential to monitor MSLN expression during tumor development.

An Oxidative Stress Nanoamplifier toEnhance the Efficacy of Cisplatin in Head and Neck Cancer.

Cisplatin (CP) is a first-line platinum-based drug used for the treatment of head and neck cancer. However, tumor cells can diminish the therapeutic effects of CP through the detoxification system mediated by glutathione (GSH) and the nucleotide excision repair (NER) pathway. Herein, we present a light-activable and pH-responsive oxidative stress nanoamplifier (FPLC@IR OSNA), comprising an amphiphilic compound (FPLC) with Fmoc-lysine acting as a connector between a nitroimidazole derivative and a pH-responsive cinnamaldehyde (CA) derivative, loaded with photosensitizer IR780. The acidic microenvironment of lysosome can trigger the release of CA to produce H2O2, which breaks down into oxygen, further improving the phototherapy efficacy mediated by IR780 irradiation. The consumption of oxygen during the phototherapy process induces hypoxia, prompting the reduction of nitroimidazole to aminoimidazole and leading to reduced GSH synthesis, enhancing tumor cell death induced by CP. Meanwhile, the accumulation of intracellular reactive oxygen species (ROS) during phototherapy attenuates the nuclear NER pathway, further augmenting the therapy effect of CP. This strategy, by combining FPLC@IR OSNA with laser and CP, significantly promotes the therapeutic effect in vitro, and notably inhibits tumor growth in both Cal27 cell line-derived xenograft models and patient-derived xenograft models.

Effective eradication of acute myeloid leukemia stem cells with FLT3-directed antibody-drug conjugates.

Refractory disease and relapse are major challenges in acute myeloid leukemia (AML) therapy attributed to survival of leukemic stem cells (LSC). To target LSCs, antibody-drug conjugates (ADCs) provide an elegant solution, combining the specificity of antibodies with highly potent payloads. We aimed to investigate if FLT3-20D9h3-ADCs delivering either the DNA-alkylator duocarmycin (DUBA) or the microtubule-toxin monomethyl auristatin F (MMAF) can eradicate quiescent LSCs. We show here that DUBA more potently kills cell-cycle arrested AML cells compared to microtubule-targeting auristatins. Due to limited stability of 20D9h3-DUBA ADC in vivo, we analyzed both ADCs in advanced in vitro stem cell assays. 20D9h3-DUBA successfully eliminated leukemic progenitors in vitro in colony-forming unit and long-term culture initiating cell assays, both in patient cells and in patient-derived xenograft (PDX) cells. Further, it completely prevented engraftment of AML PDX leukemia-initiating cells in NSG mice. 20D9h3-MMAF had a similar effect in engraftment assays, but a less prominent effect in colony assays. Both ADCs did not affect healthy stem and progenitor cells at comparable doses providing the rationale for FLT3 as therapeutic LSC target. Collectively, we show that FLT3-directed ADCs with DUBA or MMAF have potent activity against AML LSCs and represent promising candidates for further clinical development.

Receptor CDCP1 is a potential target for personalized imaging and treatment of poor outcome HER2+, triple negative and metastatic ER+/HER2- breast cancers.

Receptor CUB-domain containing- protein 1 (CDCP1) was evaluated as a target for detection and treatment of breast cancer. CDCP1 expression was assessed immunohistochemically in tumors from 423 patients (119 triple-negative breast cancer (TNBC); 75 HER2+; 229 ER+/HER2- including 228 primary tumors, 229 lymph node and 47 distant metastases). Cell cytotoxicity induced in vitro by a CDCP1-targeting antibody-drug conjugate (ADC), consisting of the human/mouse chimeric antibody ch10D7 and the microtubule disruptor monomethyl auristatin E (MMAE), was quantified, including in combination with HER2-targeting ADC T-DM1. Detection of CDCP1-expressing primary and metastatic xenografts in mice was examined by PET-CT imaging using 89zirconium-labelled ch10D7 (89Zr-ch10D7). The impact of ch10D7-MMAE on tumor burden and survival in vivo, including in combination with T-DM1, was quantified in cell line and patient-derived xenograft mouse models. CDCP1 is expressed predominantly on the surface of malignant cells of 70% of TNBCs, 80% of HER2+ tumors, and increases in ER+/HER2- tumors from 44.9% in primary tumors to 56.4% in lymph node metastases and 74.3% in distant metastases. PET-CT imaging with 89Zr-ch10D7 is effective for the detection of primary and metastatic CDCP1-expressing TNBCs in mice. ADC ch10D7-MMAE kills CDCP1-expressing cells in vitro and controls primary and metastatic TNBC xenografts in mice, conferring significant survival advantages over chemotherapy. It compares favorably to T-DM1 in vivo, and ch10D7-MMAE combined with T-DM1 showed the most potent efficacy, markedly reducing tumor burden of CDCP1+/HER2+ xenografts and prolonging mouse survival, compared with T-DM1 or ch10D7. CDCP1-directed molecular imaging has potential to identify aggressive breast cancers for CDCP1-targeted treatment.

Abstract B030: Novel strategy to improve response of high risk HPV+ HNSCC to radiation therapy

Abstract Background: Improving understanding of head and neck squamous cell carcinoma (HNSCC) has led to the discovery of novel molecular markers to stratify risk and guide treatment decisions. Recently, we have identified two subtypes of HPV+ OPSCC – one with good prognosis and one with poor prognosis. The subtypes are distinct in many ways – somatic genes mutated, HPV viral oncogenes expression, the physical state of the viral genome (integrated vs episomal), as well as global mutation and methylation profiles. Intriguingly, all differences converge on intrinsic tumor NF-κB activity with constitutively active NF-κB (usually arising from genetic defects in NF-κB regulators, including TRAF3 and CYLD) driving significantly elevated radiation sensitivity and improved patient survival. NF-κB-based groups define an important clinically actionable prognostic biomarker in HPV+ HNSCC: one important goal is to de-escalate therapy for patients with molecularly low-risk tumors to improve quality of life amongst survivors, while another goal is to improve the cure for of patients with a high risk of treatment failure. Material and methods: We utilized HPV+ head and neck cancer cells, HPV+ HNSCC xenografted mouse model, patient-derived xenografts (PDX), as well as our novel fully immunocompetent mouse model of high risk HPV+ HNSCC; we applied targeted fractionated radiation. To activate NF-κB, we used two clinically applicable drugs with different mechanisms of action: Toll-like receptor 5 (TLR5) agonist and mitochondria-derived activator of caspases (SMAC) mimetic. Custom NF-κB NanoString panel contains our recently developed NF-κB gene signature along with HPV genes and housekeeping genes that are used to normalize the gene expression data. Results: Both drugs activated NF-κB in HPV+ HNSCC and significantly improved response of HPV+ head and neck cancer cells and tumors to radiation. Interestingly, TLR5 agonist alone had a strong tumor suppressing effect in low NF-κB HPV+ HNSCC; this effect was significantly better than 32 Gy focused radiation given in fractions of 4 Gy. Custom NF-κB NanoString panel is robustly prognostic with NF-κB active tumors having improved outcomes after chemoradiation. Conclusions: Because of the average relatively favorable prognosis of HPV+ HNSCC, urgent need for improved therapy for patients with aggressive high risk tumors is greatly undervalued. Here, we present a clinical assay to risk stratify patients with HPV+ HNSCC and begin to explore pharmacological activation of NF-κB as a strategy to sensitize high risk HPV+ HNSCC to radiation therapy. TLR5 agonist is well known for the remarkable normal tissue protective effects against radiation and other damaging conditions; the dual effect of entolimod – sensitization of HPV+ HNSCC to radiation, while simultaneously protecting normal tissues from severe side effects - may be useful for patients with high-risk HPV+ head and neck cancer in combination with radiotherapy. Citation Format: Sri Vemulamanda, Natalia Issaeva1, Aditi Kothari, Travis Schrank, Wendell G. Yarbrough. Novel strategy to improve response of high risk HPV+ HNSCC to radiation therapy. [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Translating Targeted Therapies in Combination with Radiotherapy; 2025 Jan 26-29; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(2_Suppl):Abstract nr B030.

Differences in treatment outcome between translational platforms in developing therapies for gastrointestinal cancers.

The variability in translational models profoundly impacts the outcomes and predictive value of preclinical studies for gastrointestinal (GI) cancer treatments. Preclinical models, including 2D cell cultures, 3D organoids, patient-derived xenografts (PDXs), and animal models, provide distinct advantages and limitations in replicating the complex tumor microenvironment (TME) of human cancers. Each model's unique biological and structural differences contribute to discrepancies in treatment responses, challenging the direct translation of experimental results to clinical settings. While 2D cell cultures are cost-effective and suitable for high-throughput screening, they lack the 3D architecture and cellular interactions of the in vivo TME. Organoids offer a more comprehensive 3D structure that better mirrors tumor heterogeneity, yet they still face limitations in fully mimicking in vivo conditions, such as vascularization and immune cell interactions. PDXs, although more representative of human cancers due to their genetic fidelity and TME preservation, are costly and resource-intensive, with human stromal and immune components gradually replaced by murine counterparts over time. This review assesses the strengths and limitations of each model, highlighting recent advancements in translational platforms that incorporate complex TME features. Understanding the influence of model selection on treatment efficacy predictions is essential for enhancing the reliability of preclinical findings and advancing personalized therapeutic strategies for GI cancers.

Targeting the IKZF1/BCL-2 axis as a novel therapeutic strategy for treating acute T-cell lymphoblastic leukemia

ABSTRACT Objectives Acute T-cell lymphoblastic leukemia (T-ALL) is a severe hematologic malignancy with limited treatment options and poor long-term survival. This study explores the role of IKZF1 in regulating BCL-2 expression in T-ALL. Methods CUT&Tag and CUT&Run assays were employed to assess IKZF1 binding to the BCL-2 promoter. IKZF1 overexpression and knockdown experiments were performed in T-ALL cell lines. The effects of CX-4945 and venetoclax, alone and in combination, were evaluated in vitro and in vivo T-ALL models. Results CUT&Tag sequencing identified IKZF1 binding to the BCL-2 promoter, establishing it as a transcriptional repressor. Functional assays demonstrated that IKZF1 overexpression reduced BCL-2 mRNA levels and increased repressive histone marks at the BCL-2 promoter, while IKZF1 knockdown led to elevated BCL-2 expression. CX-4945, a CK2 inhibitor, could reduced BCL-2 levels in T-ALL cells. Notably, knockdown of IKZF1 partially rescued the CX-4945-induced repression of BCL-2. These results underscore the CK2-IKZF1 signaling axis as a key regulator of BCL-2 expression. In vitro, CX-4945 enhanced the cytotoxicity of venetoclax, with the combination showing significant synergistic effects and increased apoptosis in T-ALL cell lines. In vivo studies with cell line-derived xenograft (CDX) and patient-derived xenograft (PDX) models demonstrated that CX-4945 and venetoclax combined therapy provided superior therapeutic efficacy, reducing tumor burden and prolonging survival compared to single-agent treatments. Conclusions IKZF1 represses BCL-2 in T-ALL, and targeting the CK2-IKZF1 axis with CX-4945 and venetoclax offers a promising therapeutic strategy, showing enhanced efficacy and potential as a novel treatment approach for T-ALL.

Nanoparticle-Mediated mRNA Delivery to Triple-Negative Breast Cancer (TNBC) Patient-Derived Xenograft (PDX) Tumors

Nanoparticle-Mediated mRNA Delivery to Triple-Negative Breast Cancer (TNBC) Patient-Derived Xenograft (PDX) Tumors

Patient-derived xenograft models of gastrointestinal stromal tumors: a ready-to-use platform for translational research.

Gastrointestinal stromal tumors (GIST) are the most common mesenchymal malignancy of the gastrointestinal tract. Most GIST harbor mutations in oncogenes, such as KIT, and are treated with tyrosine kinase inhibitors (TKI), such as imatinib. Most tumors develop secondary mutations inducing drug resistance against the available TKI, which requires novel therapies. We established a GIST patient-derived xenograft (PDX) platform of GIST that can be used for preclinical drug testing. Tumor tissue from consenting GIST patients was transplanted subcutaneously to NMRI nu/nu mice. Once tumor growth was observed, the tumor was re-transplanted to a next generation of mice. Tumors were characterized histopathologically and molecularly at every re-transplantation and compared with the original patient tumor. We transplanted 112 tumor samples from 99 GIST patients, resulting in 12 established and well-characterized GIST models with different mutations and TKI sensitivity. Three models harbor secondary KIT mutations. One model is characterized by a primary, imatinib-resistant PDGFRA exon 18 p.D842V mutation. Our established platform of well-characterized GIST PDX models, covering the most relevant driver mutations, serves as an excellent tool for preclinical drug testing and tumor biology studies.

A surgical orthotopic xenograft approach with immune response for colorectal cancer research.

The high morbidity and mortality of colorectal cancer (CRC) is a major challenge in clinical practice. Although a series of alternative research models of CRC have been developed, appropriate orthotopic animal models that reproduce the specific clinical response as well as pathophysiological immune features of CRC are still lacking. In the current study, we constructed a CRC orthotopic xenograft model by implanting the tumor tubes at the colorectum of mice and monitored the model development using bioluminescence imaging. This model successfully recapitulates the clinical chemotherapy efficacy, including reduced total flux, tumor weight, and the expression of Ki67 after treatment of the first-line chemotherapy regime of CRC (FOLFOX: oxaliplatin and 5-fluorouracil/calcium folinate). The model also reproduced the immunosuppressive effect of FOLFOX, indicated by decreased infiltration of macrophages and increased Treg cells in tumor. Additionally, the orthotopic xenograft approach may be applied in immunodeficient NCG/NSG mice for constructing patient-derived xenografts, and being used in clinical precision medicine and drug evaluation. We believe the current model is a successful surgical orthotopic xenograft approach for cancer research and deserves to be popularized, which will provide a convenient and efficient platform for in-depth mechanism exploration of CRC and preclinical drug evaluation.

Patient-derived xenografts from circulating cancer stem cells as a preclinical model for personalized pancreatic cancer research

Patient-derived xenografts (PDXs) provide biologically relevant models and potential platforms for the development of treatment strategies for precision medicine in pancreatic cancer. Furthermore, circulating epithelial tumor cells (CETCs/CTCs) are released into the bloodstream by solid tumors and a rare subpopulation—circulating cancer stem cells (cCSCs) – is considered to be responsible for recurrence and plays a key role in metastasis. For the identification of cCSCs, an innovative in vitro assay to generate tumorspheres was established in this study. The number of tumorspheres and CETCs/CTCs was analyzed perioperatively in 25 pancreatic cancer patients. Additionally, an individual in vivo chorioallantoic membrane (CAM) culture system was used to generate PDXs from these tumorspheres. While overall correlations of CETCs/CTCs with clinicopathological parameters did not reach statistical significance, a significant difference in the number of tumorspheres was observed between patient subgroups with lower and higher UICC stages. This finding underscores their potential as biomarkers, providing valuable insights into clinical decision-making and tumor progression. The application of tumorspheres on the CAM successfully established PDXs within 7 days. These xenografts closely resembled the histological features of the primary tumor. Hence, this model represents a novel and fast option for individualized testing of new therapies for PDAC.

Targeting LMO2-induced autocrine FLT3 signaling to overcome chemoresistance in early T-cell precursor acute lymphoblastic leukemia.

Early T-cell Precursor Acute Lymphoblastic Leukemia (ETP-ALL) is an immature subtype of T-cell acute lymphoblastic leukemia (T-ALL) commonly show deregulation of the LMO2-LYL1 stem cell transcription factors, activating mutations of cytokine receptor signaling, and poor early response to intensive chemotherapy. Previously, studies of the Lmo2 transgenic mouse model of ETP-ALL identified a population of stem-like T-cell progenitors with long-term self-renewal capacity and intrinsic chemotherapy resistance linked to cellular quiescence. Here, analyses of Lmo2 transgenic mice, patient-derived xenografts, and single-cell RNA-sequencing data from primary ETP-ALL identified a rare subpopulation of leukemic stem cells expressing high levels of the cytokine receptor FLT3. Despite a highly proliferative state, these FLT3-overexpressing cells had long-term self-renewal capacity and almost complete resistance to chemotherapy. Chromatin immunoprecipitation and assay for transposase-accessible chromatin sequencing demonstrated FLT3 and its ligand may be direct targets of the LMO2 stem-cell complex. Media conditioned by Lmo2 transgenic thymocytes revealed an autocrine FLT3-dependent signaling loop that could be targeted by the FLT3 inhibitor gilteritinib. Consequently, gilteritinib impaired in vivo growth of ETP-ALL and improved the sensitivity to chemotherapy. Furthermore, gilteritinib enhanced response to the BCL2 inhibitor venetoclax, which may enable "chemo-free" treatment of ETP-ALL. Together, these data provide a cellular and molecular explanation for enhanced cytokine signaling in LMO2-driven ETP-ALL beyond activating mutations and a rationale for clinical trials of FLT3 inhibitors in ETP-ALL.

Stochastic demethylation and redundant epigenetic suppressive mechanisms generate highly heterogeneous responses to pharmacological DNA methyltransferase inhibition

BackgroundDespite promising preclinical studies, the application of DNA methyltransferase inhibitors in treating patients with solid cancers has thus far produced only modest outcomes. The presence of intratumoral heterogeneity in response to DNA methyltransferase inhibitors could significantly influence clinical efficacy, yet our understanding of the single-cell response to these drugs in solid tumors remains very limited.MethodsIn this study, we used cancer/testis antigen genes as a model for methylation-dependent gene expression to examine the activity of DNA methyltransferase inhibitors and their potential synergistic effect with histone deacetylase inhibitors at the single-cancer cell level. The analysis was performed on breast cancer patient-derived xenograft tumors and cell lines, employing a comprehensive set of techniques, including targeted single-cell mRNA sequencing. Mechanistic insights were further gained through DNA methylation profiling and chromatin structure analysis.ResultsWe show that breast cancer tumors and cell cultures exhibit a highly heterogenous response to DNA methyltransferase inhibitors, persisting even under high drug concentrations and efficient DNA methyltransferase depletion. The observed variability in response to DNA methyltransferase inhibitors was independent of cancer-associated aberrations and clonal genetic diversity. Instead, these variations were attributed to stochastic demethylation of regulatory CpG sites and the DNA methylation-independent suppressive function of histone deacetylases.ConclusionsOur findings point to intratumoral heterogeneity as a limiting factor in the use of DNA methyltransferase inhibitors as single agents in treatment of solid cancers and highlight histone deacetylase inhibitors as essential partners to DNA methyltransferase inhibitors in the clinic.