Long-term Tumor Growth Research Articles

Ultrasound Molecular Imaging Enhances High-Intensity Focused Ultrasound Ablation on Liver Cancer With B7-H3-Targeted Microbubbles.

High-intensity focused ultrasound (HIFU) is a promising minimally invasive treatment for liver cancer; however, its efficacy is often limited by the attenuation of ultrasonic energy. This study investigates the effectiveness of B7-H3-targeted microbubbles (T-MBs) in enhancing HIFU ablation of liver cancer and explores their potential for clinical translation. T-MBs and isotype control microbubbles (I-MBs) were synthesized through the conjugation of biotinylated anti-B7-H3 antibody and isotype control antibody to the microbubble surface, respectively. Contrast-enhanced ultrasound imaging was performed to compare the accumulation of T-MBs and I-MBs in liver cancer at various time points. The efficacy of T-MBs in enhancing HIFU treatment was evaluated by measuring the immediate tumor ablation rate and long-term tumor growth suppression. Additionally, the induced antitumor immune response was assessed through cytokine quantification in serum and tumor tissue, along with immunofluorescence staining conducted on days 1, 3, and 7 post-treatment. T-MBs demonstrated superior liver cancer-specific accumulation, characterized by higher concentrations and prolonged retention compared to I-MBs. The combination of T-MBs with HIFU resulted in significantly enhanced tumor ablation rates and superior tumor growth suppression. Post-treatment analysis revealed a gradual uptick in cytokine levels within the tumor microenvironment, along with progressive infiltration of antitumor immune cells. T-MBs effectively enhance the therapeutic efficacy of HIFU for liver cancer treatment while simultaneously promoting an antitumor immune response. These findings provide a strong experimental foundation for the clinical translation of ultrasound molecular imaging combined with HIFU as a novel approach for tumor therapy.

TrxT and dhd are dispensable for Drosophila brain development but essential for l(3)mbt brain tumour growth

Expression of the Drosophila cancer-germline (CG), X-linked, head-to-head gene pair TrxT and dhd is normally germline-specific but becomes upregulated in brain tumours caused by mutation in l(3)mbt. Here, we show that TrxT and dhd play a major synergistic role in the emergence of l(3)mbt tumour-linked transcriptomic signatures and tumour development, which is remarkable, taking into account that these two genes are never expressed together under normal conditions. We also show that TrxT, but not dhd, is crucial for the growth of l(3)mbt allografts, hence suggesting that the initial stages of tumour development and long-term tumour growth may depend on different molecular pathways. In humans, head-to-head inverted gene pairs are abundant among CG genes that map to the X chromosome. Our results identify a first example of an X-linked, head-to-head CG gene pair in Drosophila, underpinning the potential of such CG genes, dispensable for normal development and homoeostasis of somatic tissue, as targets to curtail malignant growth with minimal impact on overall health.

Abstract PS12-03: p53 loss enables HR+ breast cancer escape from CDK4/6 inhibitor-induced quiescence via CDK2

Abstract Background: Inhibition of CDK4/6 kinases has led to improved clinical outcomes in hormone receptor positive (HR+) breast cancer. While these are highly effective therapies, only a minority of patients experience long-term disease control. We sought to determine the genomic configurations and underlying mechanisms associated with long-term response. Methods: To identify genomic patterns associated with clinical outcomes, we analyzed a cohort of 447 patients with metastatic HR+ breast cancer treated at MSK with first-line CDK4/6 inhibitors (CDK4/6i) for which tumor-normal sequencing and long-term clinical follow up were available. To identify the pattern of genomic features associated with longer, intermediate, and short response, we implemented an elastic net Cox regression on binary pathogenic variant status of each gene as well as select clinical features (prior endocrine therapy, endocrine therapy partner, de novo metastatic status). Our principal aim was variable interpretability over pure predictive accuracy. Human HR+ breast cancer models including human breast cancer organoids and cell lines were utilized for mechanistic studies. For validation in a clinical setting, we analyzed the association between Ki67 score after neoadjuvant ribociclib plus endocrine therapy and pre-treatment gene mutation from the FELINE trial [NCT02712723] using a Fisher exact test. Results: Our model identified a “longer response” group (n = 124, 27.7%) from patients with a median progression free survival (PFS) of 32.5 months, compared with an “intermediate” (n = 224, 50.1%, median PFS = 13.7 months) and “short response” group (n = 99, 22.1%, median PFS = 5.84 months). TP53 and MDM2 pathogenic variant status were the most important variables to stratify between these groups, obtaining variable selection frequencies of 1.0 and 0.93 and mean hazard ratios of 2.02 and 1.38, respectively. To elucidate the mechanisms whereby the p53 pathway supports long term response, we generated isogenic and patient derived models of TP53 loss or MDM2 overexpression. Using immunoblotting and cell cycle assays, we found that drug-treated p53 KO cells and MDM2 overexpressing cells effectively suppressed RB1 phosphorylation and blocked in G1 after 24-48 hours. However, upon drug withdrawal, these cells could reenter the cell cycle and promote long-term tumor outgrowth. These effects we observed both in vitro and in vivo. Measures of long-term CDK4/6i response such as expression of senescence associated secretory phenotype genes was abrogated by TP53 loss. Mechanistically, we found persistent phosphorylation of the p130 RB1-like protein in the p53 KO cells. Phosphorylation of p130 impaired its interaction with E2F4, thereby blocking DREAM complex assembly and promoting cell cycle reentry. Inhibition of phosphorylation of p130 via p21 overexpression or by selective CDK2 inhibitors could restore irreversible cell cycle arrest in p53 KO cells. The combination of CDK2 and CDK4/6 inhibition led to long-term tumor growth suppression in models with mutant TP53. To validate the human relevance of TP53 mediating CDK4/6i response, we analyzed longitudinal samples from the FELINE trial that evaluated efficacy and feasibility of neoadjuvant ribociclib plus endocrine therapy. Of 45 evaluable patients, 13 (28.9%) harbored a pre-treatment TP53 loss of function variant. Of these 13 cases, 7 (53.8%) did not achieve a low ( < 10%) Ki-67 upon surgical resection as compared to TP53 wildtype tumors (n=32), only one (3.2%) of which did not achieve a low Ki-67 [OR 32.1, 95% CI 3.28 – 1660.3, p = 0.00026]. Conclusion: Loss of p53 was strongly associated with lack of long-term response to CDK4/6i in patients. Complete inhibition of both CDK4/6 and CDK2 appears to be necessary in order to convert quiescent HR+ tumors cells into durably inhibited and effectively dormant cancers. Citation Format: Rei Kudo, Anton Safonov, Edaise M. da Silva, Qing Li, Hong Shao, Marie Will, Harikrishna Nakshatri, Jorge Reis-Filho, Shom Goel, Andrew Koff, Britta Weigelt, Qamar Khan, Pedram Razavi, Sarat Chandarlapaty. p53 loss enables HR+ breast cancer escape from CDK4/6 inhibitor-induced quiescence via CDK2 [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PS12-03.

Combination of BET Inhibition with Radiation Induces Anti-Tumor Immune Response in Pre-Clinical Models of ER-Positive Breast Cancer

Despite advances in multidisciplinary care, the risk of late recurrence in Estrogen Receptor-positive (ER+) breast cancer (BC) patients following standard-of-care treatment ranges from 20-52% at 20 years. While immune checkpoint inhibitors (CPI), have shown enormous potential in inducing long term remission in some cases, patients with ER+ BC show poor response to these agents. The objective of this study is to develop effective radiation therapy (RT)-based combination regimens that induce anti-tumor immunity against immunologically cold ER+ breast tumors. We have previously shown that pharmacological inhibition of bromodomain and extraterminal domain (BET) family of epigenetic regulator proteins overcomes both endocrine therapy resistance and radiation resistance in pre-clinical models of ER+ BC by reversing pathological reprogramming of transcriptional and DNA repair pathways, respectively. In this study, we explore the potential of RT+BET inhibition combination to elicit anti-tumor immunity against ER+ BC. A unique ER+ BC model, MXT+, was used to generate tumors in female, BDF1 mice. PD-L1 expression following RT or RT+OTX015 (a BET inhibitor) was assessed by Western blot analysis. Comet assay was used to assess the degree of unrepaired DNA damage following treatment with RT alone, OTX015 alone and RT+OTX015. Multiplex IHC was used to assess changes in tumor microenvironment after various treatments. Tumor growth characteristics were established following treatment with vehicle, RT alone (8 Gy x 3), OTX015 alone (100 mg/kg x 3 days) and RT+OTX015. Using murine ER+ BC model, MXT+, we show that RT is a potent inducer of PD-L1 expression. Combination of RT+OTX015 completely ablated RT-mediated PD-L1 induction at the transcription level. OTX015 also accentuated RT-induced DNA damage through inhibition of NHEJ pathway, resulting in increased micronuclei formation (which activate cGAS/STING pathway, a critical bridge between innate and adaptive immunity). Combination of OTX015 with RT also caused a statistically significant (on unpaired t test) increase tumor infiltrating lymphocytes. In in vivo therapeutic studies, combination of OTX015 with RT (8 Gy x 3) resulted in extraordinary synergy and caused complete inhibition of long-term tumor growth. Treatment with RT alone (8 Gy x 3) or OTX015 alone (on 3 consecutive days) only caused modest inhibition of tumor growth. ANOVA with Dunnett's test was used to adjust for multiple comparisons and showed statistically significant difference in tumor growth between OTX015+RT arm and OTX alone as well as OTX015+RT arm and RT alone. Patients with ER+ BC face high risk of late recurrence after definitive treatment and show poor response to immune CPIs. We show that combination of BET inhibition with RT induces anti-tumor immunity and completely prevents tumor growth. Clinical validation of this regimen is warranted to prevent late recurrences in ER+ BC patients.

Generation of Orthotopic Patient-Derived Xenografts in Humanized Mice for Evaluation of Emerging Targeted Therapies and Immunotherapy Combinations for Melanoma.

Current methodologies for developing PDX in humanized mice in preclinical trials with immune-based therapies are limited by GVHD. Here, we compared two approaches for establishing PDX tumors in humanized mice: (1) PDX are first established in immune-deficient mice; or (2) PDX are initially established in humanized mice; then established PDX are transplanted to a larger cohort of humanized mice for preclinical trials. With the first approach, there was rapid wasting of PDX-bearing humanized mice with high levels of activated T cells in the circulation and organs, indicating immune-mediated toxicity. In contrast, with the second approach, toxicity was less of an issue and long-term human melanoma tumor growth and maintenance of human chimerism was achieved. Preclinical trials from the second approach revealed that rigosertib, but not anti-PD-1, increased CD8/CD4 T cell ratios in spleen and blood and inhibited PDX tumor growth. Resistance to anti-PD-1 was associated with PDX tumors established from tumors with limited CD8+ T cell content. Our findings suggest that it is essential to carefully manage immune editing by first establishing PDX tumors in humanized mice before expanding PDX tumors into a larger cohort of humanized mice to evaluate therapy response.

Impact of Postoperative Stereotactic Body Radiation Therapy on Survival of Patients with Spinal Metastases in the Context of Additional Systemic Adjuvant Therapy

Stereotactic body radiotherapy (SBRT) has been established as a safe and effective treatment modality for control of long-term pain and tumor growth. However, few studies have investigated the efficacy of postoperative SBRT versus conventional external beam radiation therapy (EBRT) in extending survival within the context of systemic therapy. A retrospective chart review of patients who underwent surgery for spinal metastasis at our institution was conducted. Demographic, treatment and outcome data were collected. SBRT was compared with EBRT and non-SBRT, and analyses were stratified by whether or not patients received systemic therapy. Survival analysis was conducted using propensity score matching. Bivariate analysis in the non-systemic therapy group revealed longer survival with SBRT compared to EBRT and non-SBRT. Further analysis also showed that primary cancer type and preoperative mRS significantly affected survival. Within patients who received systemic therapy, overall median survival for SBRT patients was 22.7 months (95% CI: 12.1 - 52.3) versus 16.1 months (95% CI: 12.7 - 44.0; p = 0.28) for EBRT patients and 16.1 months (95% CI: 12.2 - 21.9; p = 0.07) for non-SBRT patients. Within patients who did not receive systemic therapy, overall median survival for SBRT patients was 62.1 months (95% CI: 18.1 - unknown) versus 5.3 months (95% CI: 2.8 - unknown; p = 0.08) for EBRT patients and 6.9 months (95% CI: 5.0 - 45.6; p = 0.02) for non-SBRT patients. In patients who do not receive systemic therapy, treatment with postoperative SBRT may increase survival time compared to patients not receiving SBRT.

Long-term tumor growth and hearing after conservative management of vestibular schwannomas

Background There has been no report of a detailed long-term study of hearing at defined frequencies. Aims/Objectives This study aimed to evaluate long-term vestibular shwannoma (VS) growth and long-term changes in hearing thresholds at defined frequencies. Methods We retrospectively retrieved the medical records of 67 VS patients. Cases that were followed up for more than 5 years were analyzed. Results Tumor growth was observed in 15 cases (22.4%) and 6 cases underwent gamma knife treatment (9.0%). The longest case of tumor growth and gamma knife treatment was observed 11 years after the initial diagnosis. Hearing thresholds at 500–2000 Hz was significantly different between the no growth and growth group (p < .05). Particularly at 1000 Hz, there was a significant difference between the two groups from an early time-point. Conclusions and Significance Even a 10-year or longer follow-up of VS may show an increase in tumor growth. Moreover, hearing thresholds, particularly at 1000 kHz, may predict tumor growth when following-up VS patients.

Evaluating the safety of perioperative dexamethasone treatment: A retrospective analysis of a single center pediatric low-grade glioma cohort.

In addition to surgical management, corticosteroids have proven to be beneficial in the management of acute symptoms related to CNS tumors, and have been widely used for many decades, with dexamethasone (DM) representing the most commonly used agent. However, lately published in vitro data possibly indicates a DM-induced suppression of oncogene-induced senescence (OIS) in a preclinical pediatric low-grade glioma (pLGG) model, which, alongside data associating perioperative DM treatment with reduced event-free survival in adult glioma, raises questions concerning the safety of DM treatment in pLGG. A total of 172 patients with pLGG were retrospectively analyzed concerning the impact of perioperative DM application on postoperative short- and long-term tumor growth velocity and progression-free survival (PFS). Three-dimensional volumetric analyses of sequential MRI follow-up examinations were used for assessment of tumor growth behavior. Mean follow-up period accounted for 60.1months. Sixty-five patients (45%) were perioperatively treated with DM in commonly used doses. Five-year PFS accounted for 93% following gross-total resection (GTR) and 57% post incomplete resection (IR). Comparison of short- and long-term postoperative tumor growth rates in patients with vs without perioperative DM application showed no significant difference (short-term: 0.022 vs 0.023 cm3 /month, respectively; long-term: 0.019 vs 0.023 cm3 /month, respectively). Comparison of PFS post IR (5-year-PFS: 65% vs 55%, respectively; 10-year-PFS: 52% vs 53%, respectively) and GTR (5- and 10-years-PFS: 91% vs 92%, respectively) likewise showed similarity. This data emphasizes the safety of perioperative DM application in pLGG, adding further evidence for decision making and requested future guidelines.

Irreversible electroporation: present and future in the treatment of hepatocellular carcinoma

Currently, among the possible treatments for hepatocellular carcinoma there is group of minimally invasive ablation techniques with wide clinical ac-ceptance due to their greater efficacy and safety in comparison to traditional therapies, low cost, and no need of being admitted to hospital (outpatient treatment program). Irreversible electroporation is a non-thermal abla-tion technique in which electrical fields are used to create nanopores in the cell membrane that induce tumor cell death. Irreversible electroporation has shown promising results in numerous clinical trials; however, its control on long-term tumor growth and recurrence is inferior in comparison to that of radiofrequency. Combining irreversible electroporation with immunological agents may increase its efficacy in the treatment of focal lesions and metastases. In this work, we present an update on IRE including procedure, mechanism of action, application as a treatment for HCC, and the improvements that have been made in the past few years.

Growth dynamics of incidental meningiomas: A prospective long-term follow-up study.

There is no consensus on the management of incidental meningiomas. The literature on long-term growth dynamics is sparse and the natural history of these tumors remains to be illuminated. We prospectively assessed long-term tumor growth dynamics and survival rates during active monitoring of 62 patients (45 female, mean age 63.9 years) harboring 68 tumors. Clinical and radiological data were obtained every 6 months for 2 years, annually until 5 years, then every second year. The natural progression of incidental meningiomas during 12 years of monitoring was growth (P < .001). However, mean growth decelerated at 1.5 years and became insignificant after 8 years. Self-limiting growth patterns were seen in 43 (63.2%) tumors, non-decelerating in 20 (29.4%) and 5 (7.4%) were inconclusive due to ≤ 2 measurements. Decelerating growth persisted once established. Within 5 years, 38 (97.4%) of 39 interventions were initiated. None developed symptoms prior to intervention. Large tumors (P < .001) involving venous sinuses (P = .039) grew most aggressively. Since inclusion 19 (30.6%) patients have died of unrelated causes and 2 (3%) from grade 2 meningiomas. Active monitoring seems a safe and appropriate first-line management of incidental meningiomas. Intervention was avoided in > 40% with indolent tumors in this cohort. Treatment was not compromised by tumor growth. Clinical follow-up seems sufficient beyond 5 years if self-limiting growth is established. Steady or accelerating growth warrant monitoring until they reach a stable state or intervention is initiated.

Early functional mismatch between breast cancer cells and their tumour microenvironment suppresses long term growth

Cancer cells thrive when embedded in a fine-tuned cellular and extracellular environment or tumour microenvironment (TME). There is a general understanding of a co-evolution between cancer cells and their surrounding TME, pointing at a functional connection between cancer cells characteristics and the perturbations induced in their surrounding tissue. However, it has never been formally proven whether this functional connection needs to be set from the start or if aggressive cancer cells always dominate their microenvironmental any point in time. This would require a dedicated experimental setting where malignant cells are challenged to grow in a different TME from the one they would naturally create. Here we generated an experimental setting where we transiently perturb the secretory profile of aggressive breast cancer cells without affecting their intrinsic growth ability, which led to the initial establishment of an atypical TME. Interestingly, even if initially tumours are formed, this atypical TME evolves to impair long term in vivo cancer growth. Using a combination of in vivo transcriptomics, protein arrays and in vitro co-cultures, we found that the atypical TME culminates in the infiltration of macrophages with STAT1high activity. These macrophages show strong anti-tumoural functions which reduce long-term tumour growth, despite lacking canonical M1 markers. Importantly, gene signatures of the mesenchymal compartment of the TME, as well as the anti-tumoural macrophages, show striking prognostic power that correlates with less aggressive human breast cancers.

Type-I interferon signaling is essential for robust metronomic chemo-immunogenic tumor regression in murine breast cancer.

Many patients with breast cancer have a poor prognosis with limited therapeutic options. Here, we investigated the potential of chemo-immunogenic therapy as an avenue of treatment. We utilized two syngeneic mouse mammary tumor models, 4T1 and E0771, to examine the chemo-immunogenic potential of cyclophosphamide and the mechanistic contributions of cyclophosphamide-activated type-I interferon (IFN) signaling to therapeutic activity. Chemically-activated cyclophosphamide induced robust IFNα/β receptor-1-dependent signaling linked to hundreds of IFN-stimulated gene responses in both cell lines. Further, in 4T1 tumors, cyclophosphamide given on a medium-dose, 6-day intermittent metronomic schedule induced strong IFN signaling but comparatively weak immune cell infiltration associated with long-term tumor growth stasis. Induction of IFN signaling was somewhat weaker in E0771 tumors but was followed by widespread downstream gene responses, robust immune cell infiltration and extensive, prolonged tumor regression. The immune dependence of these effective anti-tumor responses was established by CD8 T-cell immunodepletion, which blocked cyclophosphamide-induced E0771 tumor regression and led to tumor stasis followed by regrowth. Strikingly, IFNα/β receptor-1 antibody blockade was even more effective in preventing E0771 immune cell infiltration and blocked the major tumor regression induced by cyclophosphamide treatment. Type-I IFN signaling is thus essential for the robust chemo-immunogenic response of these tumors to cyclophosphamide administered on a metronomic schedule.

Stereotactic radiosurgery for vestibular schwannoma: early and long-term radiation-induced changes and tumor growth control

To investigate the changes of vestibular schwannoma in different periods after radiosurgery. We analyzed early and long-term radiation-induced changes in 333 patients who received Gamma Knife treatment at the Burdenko Neurosurgery Center between April 2005 and December 2015. Mean follow-up period was 60 months (range 15-167). There were 89 men (26.7%) and 244 (73.4%) women. Mean age of patients was 48.2 years. Mean baseline tumor volume was 4.1 cm3 (range 0.1-14.5). Dynamics of changes was assessed using volumetric comparison. Tumor shrinkage without pseudo-progression was observed in 149 (44.7%) patients. Typical pseudo-progression in different variants was found in 131 (39.3%) patients, i.e. short-term (1 year) and long-term (≥2 years) course, complete and incomplete process. Eleven patients had atypical pseudo-progression after initial tumor shrinkage. Progression-free 5- and 10-year survival in the entire group was 87 and 81%, respectively. Progression-free 5-year survival rate was 95 and 92% in patients with and without pseudo-progression, respectively. Ten-year survival rate was 89 and 89%, respectively. Knowledge of pseudo-progression features is essential for the most reasonable and reliable assessment of treatment results and justification of timing and frequency of subsequent MR control. Atypical course of pseudo-progression can simulate tumor recurrence. In case of tumor enlargement at any follow-up stage after radiosurgery, advisability of surgery should be determined considering clinical data and likelihood of tumor shrinkage following natural regression of post-radiation tumor enlargement.

Imaging-Guided Evaluation of the Novel Small-Molecule Benzosuberene Tubulin-Binding Agent KGP265 as a Potential Therapeutic Agent for Cancer Treatment.

Simple SummaryVascular-disrupting agents promise significant therapeutic efficacy against solid tumors by selectively damaging tumor-associated vasculature. Dynamic BLI and oxygen-enhanced multispectral optoacoustic tomography (OE-MSOT) were used to compare vascular shutdown following administration of KGP265. BLI signal and vascular oxygenation response (ΔsO2) to a gas breathing challenge were both significantly reduced within 2 h indicating vascular disruption, which continued over 24 h. Twice-weekly doses of KGP265 caused a significant growth delay in MDA-MB-231 human breast tumor xenografts and 4T1 syngeneic breast tumors growing orthotopically in mice.The selective disruption of tumor-associated vasculature represents an attractive therapeutic approach. We have undertaken the first in vivo evaluation of KGP265, a water-soluble prodrug of a benzosuberene-based tubulin-binding agent, and found promising vascular-disrupting activity in three distinct tumor types. Dose escalation in orthotopic MDA-MB-231-luc breast tumor xenografts in mice indicated that higher doses produced more effective vascular shutdown, as revealed by dynamic bioluminescence imaging (BLI). In syngeneic orthotopic 4T1-luc breast and RENCA-luc kidney tumors, dynamic BLI and oxygen enhanced multispectral optoacoustic tomography (OE-MSOT) were used to compare vascular shutdown following the administration of KGP265 (7.5 mg/kg). The BLI signal and vascular oxygenation response (ΔsO2) to a gas breathing challenge were both significantly reduced within 2 h, indicating vascular disruption, which continued over 24 h. A correlative histology confirmed increased necrosis and hemorrhage. Twice-weekly doses of KGP265 caused significant growth delay in both MDA-MB-231 and 4T1 breast tumors, with no obvious systemic toxicity. A combination with carboplatin produced significantly greater tumor growth delay than carboplatin alone, though significant carboplatin-associated toxicity was observed (whole-body weight loss). KGP265 was found to be effective at low concentrations, generating long-term vascular shutdown and tumor growth delay, thus providing strong rationale for further development, particularly in combination therapies.

Association of immune-related pneumonitis with clinical benefit of anti-programmed cell death-1 monotherapy in advanced non-small cell lung cancer.

BackgroundThe association between the development of checkpoint inhibitor pneumonitis (CIP) with tumor response and survival has remained unclear so far. The aim of the present study was to evaluate the association between CIP and the clinical efficacy of anti‐programmed cell death‐1 antibody in patients with advanced non‐small cell lung cancer (NSCLC).MethodsBetween January 2016 and August 2019, 203 advanced NSCLC patients were administered with nivolumab or pembrolizumab. Comparisons were made between patients with and without CIP. We evaluated the time‐to‐treatment failure (TTF), progression‐free survival (PFS), and overall survival (OS).ResultsCIP was observed in 28 (14%) patients. CIP was associated with a longer PFS (18.9 months [95% confidence interval, CI: 8.7 months–not reached] vs. 3.9 months [95% CI: 3.4–5.1 months, p < 0.01]) and longer OS (27.4 [95% CI: 20.7 months–not reached] vs. 14.8 months [95% CI: 11.2–17.9 months, p = 0.003]). Most patients discontinued the immune checkpoint inhibitor (ICI) treatment when they developed CIP. Seven patients (25%) lived for more than 300 days from treatment discontinuation and did not show any long‐term tumor growth after treatment discontinuation.ConclusionCIP was associated with prolonged PFS and OS. Additionally, 25% of CIP patients did not show any tumor growth for long periods after treatment discontinuation. Careful management of CIP can help in obtaining the best clinical efficacy from anti‐PD‐1 antibody.

Combined Gemcitabine and Immune-Checkpoint Inhibition Conquers Anti-PD-L1 Resistance in Low-Immunogenic Mismatch Repair-Deficient Tumors.

Tumors arising in the context of Lynch Syndrome or constitutional mismatch repair deficiency are hypermutated and have a good response towards immune-checkpoint inhibitors (ICIs), including α-PD-L1 antibodies. However, in most cases, resistance mechanisms evolve. To improve outcomes and prevent resistance development, combination approaches are warranted. Herein, we applied a combined regimen with an α-PD-L1 antibody and gemcitabine in a preclinical tumor model to activate endogenous antitumor immune responses. Mlh1−/− mice with established gastrointestinal tumors received the α-PD-L1 antibody (clone 6E11; 2.5 mg/kg bw, i.v., q2wx3) and gemcitabine (100 mg/kg bw, i.p., q4wx3) in mono- or combination therapy. Survival and tumor growth were recorded. Immunological changes in the blood were routinely examined via multi-color flow cytometry and complemented by ex vivo frameshift mutation analysis to identify alterations in Mlh1−/−-tumor-associated target genes. The combined therapy of α-PD-L1 and gemcitabine prolonged median overall survival of Mlh1−/− mice from four weeks in the untreated control group to 12 weeks, accompanied by therapy-induced tumor growth inhibition, as measured by [18F]-FDG PET/CT. Plasma cytokine levels of IL13, TNFα, and MIP1β were increased and also higher than in mice receiving either monotherapy. Circulating splenic and intratumoral myeloid-derived suppressor cells (MDSCs), as well as M2 macrophages, were markedly reduced. Besides, residual tumor specimens from combi-treated mice had increased numbers of infiltrating cytotoxic T-cells. Frameshift mutations in APC, Tmem60, and Casc3 were no longer detectable upon treatment, likely because of the successful eradication of single mutated cell clones. By contrast, novel mutations appeared. Collectively, we herein confirm the safe application of combined chemo-immunotherapy by long-term tumor growth control to prevent the development of resistance mechanisms.

Preclinical and Clinical Evidence of Immune Responses Triggered in Oncologic Photodynamic Therapy: Clinical Recommendations.

Photodynamic therapy (PDT) is an anticancer strategy utilizing light-mediated activation of a photosensitizer (PS) which has accumulated in tumor and/or surrounding vasculature. Upon activation, the PS mediates tumor destruction through the generation of reactive oxygen species and tumor-associated vasculature damage, generally resulting in high tumor cure rates. In addition, a PDT-induced immune response against the tumor has been documented in several studies. However, some contradictory results have been reported as well. With the aim of improving the understanding and awareness of the immunological events triggered by PDT, this review focuses on the immunological effects post-PDT, described in preclinical and clinical studies. The reviewed preclinical evidence indicates that PDT is able to elicit a local inflammatory response in the treated site, which can develop into systemic antitumor immunity, providing long-term tumor growth control. Nevertheless, this aspect of PDT has barely been explored in clinical studies. It is clear that further understanding of these events can impact the design of more potent PDT treatments. Based on the available preclinical knowledge, recommendations are given to guide future clinical research to gain valuable information on the immune response induced by PDT. Such insights directly obtained from cancer patients can only improve the success of PDT treatment, either alone or in combination with immunomodulatory approaches.

Establishing a cryopreservation protocol for patient-derived xenografts of prostate cancer.

Serially transplantable patient-derived xenografts (PDXs) are invaluable preclinical models for studying tumor biology and evaluating therapeutic agents. As these models are challenging to establish from prostate cancer specimens, the ability to preserve them through cryopreservation has several advantages for ongoing research. Despite this, there is still uncertainty about the ability to cryopreserve PDXs of prostate cancer. This study compared three different cryopreservation protocols to identify a method that can be used to reproducibly cryopreserve a diverse cohort of prostate cancer PDX models. One serially transplantable prostate cancer PDX from the Melbourne Urological Research Alliance cohort was used to compare three cryopreservation protocols: slow freezing in fetal calf serum (FCS) with 10% dimethyl sulfoxide (DMSO), FCS with 10% DMSO supplemented with the Rho-associated kinase (ROCK) inhibitor Y-27632 and vitrification. The efficiency of the slow freezing protocols was then assessed in 17 additional prostate cancer PDXs. Following cryopreservation, PDXs were re-established in host mice that were either intact and supplemented with testosterone or castrated. Graft take rate, tumor growth, histological features, and transcriptome profiles before and after cryopreservation were compared. Slow freezing maintained the viability and histological features of prostate cancer PDXs, and the addition of a ROCK inhibitor increased their growth following cryopreservation. Using the slow freezing method, we re-established 100% of PDXs grown in either testosterone-supplemented or castrated host mice. Importantly, the long-term tumor growth rate and transcriptome profile were maintained following cryopreservation. This study has identified a protocol to reliably cryopreserve and re-establish a diverse cohort of serially transplantable PDXs of prostate cancer. This study has the potential to significantly improve the practicality of maintaining PDX models. Cryopreservation may also increase the accessibility of these important resources and provide new opportunities for preclinical studies on a broader spectrum of prostate tumors.

Modeling long-term tumor growth and kill after combinations of radiation and radiosensitizing agents

PurposeRadiation therapy, whether given alone or in combination with chemical agents, is one of the cornerstones of oncology. We develop a quantitative model that describes tumor growth during and after treatment with radiation and radiosensitizing agents. The model also describes long-term treatment effects including tumor regrowth and eradication.MethodsWe challenge the model with data from a xenograft study using a clinically relevant administration schedule and use a mixed-effects approach for model-fitting. We use the calibrated model to predict exposure combinations that result in tumor eradication using Tumor Static Exposure (TSE).ResultsThe model is able to adequately describe data from all treatment groups, with the parameter estimates taking biologically reasonable values. Using TSE, we predict the total radiation dose necessary for tumor eradication to be 110 Gy, which is reduced to 80 or 30 Gy with co-administration of 25 or 100 mg kg−1 of a radiosensitizer. TSE is also explored via a heat map of different growth and shrinkage rates. Finally, we discuss the translational potential of the model and TSE concept to humans.ConclusionsThe new model is capable of describing different tumor dynamics including tumor eradication and tumor regrowth with different rates, and can be calibrated using data from standard xenograft experiments. TSE and related concepts can be used to predict tumor shrinkage and eradication, and have the potential to guide new experiments and support translations from animals to humans.

Targeting the TLK1/NEK1 DDR axis with Thioridazine suppresses outgrowth of androgen independent prostate tumors.

Standard therapy for advanced Prostate Cancer (PCa) consists of antiandrogens, which provide respite from disease progression, but ultimately fail resulting in the incurable phase of the disease: mCRPC. Targeting PCa cells before their progression to mCRPC would greatly improve the outcome. Combination therapy targeting the DNA Damage Response (DDR) has been limited by general toxicity, and a goal of clinical trials is how to target the DDR more specifically. We now show that androgen deprivation therapy (ADT) of LNCaP cells results in increased expression of TLK1B, a key kinase upstream of NEK1 and ATR and mediating the DDR that typically results in a temporary cell cycle arrest of androgen responsive PCa cells. Following DNA damage, addition of the TLK specific inhibitor, thioridazine (THD), impairs ATR and Chk1 activation, establishing the existence of a ADT > TLK1 > NEK1 > ATR > Chk1, DDR pathway, while its abrogation leads to apoptosis. Treatment with THD suppressed the outgrowth of androgen‐independent (AI) colonies of LNCaP and TRAMP‐C2 cells cultured with bicalutamide. Moreover, THD significantly inhibited the growth of several PCa cells in vitro (including AI lines). Administration of THD or bicalutamide was not effective at inhibiting long‐term tumor growth of LNCaP xenografts. In contrast, combination therapy remarkably inhibited tumor growth via bypass of the DDR. Moreover, xenografts of LNCaP cells overexpressing a NEK1‐T141A mutant were durably suppressed with bicalutamide. Collectively, these results suggest that targeting the TLK1/NEK1 axis might be a novel therapy for PCa in combination with standard of care (ADT).