Adaptation of a Xenograft AML Model to Evaluate Chemotherapeutic Efficacy In Vivo

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3304-3304 ◽  
Author(s):  
Mark Wunderlich ◽  
Fu-Sheng Chou ◽  
Mahesh Shrestha ◽  
Benjamin Mizukawa ◽  
James C. Mulloy
Keyword(s):  
Blood Cell ◽  
Treatment Period ◽  
Conflicts Of Interest ◽  
Residual Disease ◽  
Synergistic Effects ◽  
Day Treatment ◽  
Chemotherapeutic Agents ◽  
Mouse Strains

Abstract Abstract 3304 Although significant progress has been made in the treatment of leukemia, relapse continues to be a major problem, particularly in acute myeloid leukemia (AML). The prognosis for relapsed leukemia is poor, indicating an area for potential improvements. However, animal models to study the response of human AML to chemotherapeutics and subsequent relapse are lacking. Recently we developed an improved NOD/SCID mouse with IL2RG knockout and transgenic expression of myelo-supportive cytokines SCF, GM-CSF, and IL-3 (the NSGS mouse). This mouse is remarkable in its ability to accept human AML grafts more efficiently than all other available strains. When coupled with in vitro derived AML cells, the NSGS mouse allows for a more predictable AML model with shorter latency and smaller range of death than in other mouse strains, including NSG mice. Importantly, very low numbers of cells reliably generate fatal AML in roughly 40 days, even in non-irradiated NSGS mice, allowing for rapid experimental conclusions and reduced toxicity. With the benefits of these unique tools, we sought to develop a model system to evaluate the efficacy of chemotherapeutic agents on human AML cells in vivo. Engrafted mice received a chemotherapy regimen over a 5-day treatment period consisting of a daily dose of cytarabine with simultaneous injection of doxorubicin during the first three days. Treated mice experienced striking weight loss during the treatment period with a nadir at days 8–10 post-treatment. Mice recovered body weight within 3 weeks. Serial complete blood counts indicated a rapid transient drop in total white blood cell and neutrophil counts and a delayed transient drop in red blood cell and platelet numbers, reminiscent of the effects observed in patients undergoing chemotherapy. The drugs successfully targeted the cells of the bone marrow, as evidenced by a profound loss of cellularity in treated mice relative to controls. When mice harboring N-Ras(G12D) positive AML cells were treated at early time points post-transplant, a significant reduction of tumor burden was observed in the BM and PB, with the grafts of treated mice essentially undetectable for weeks after treatment cessation. Nevertheless, treated mice inevitably succumbed to disease, although with a significantly prolonged latency compared to mock treated mice. However, when AML cells containing the FLT3-ITD mutation were used, a shift in disease latency was not reproducibly seen. This data correlates well with patient data showing that FLT3-ITD mutant AML has a worse prognosis than AML samples with N-Ras mutations. Importantly, the reappearance of AML within weeks of treatment affords the opportunity to model drug resistance and relapse, as well as the potential synergistic effects of experimental compounds used in combination with traditional chemotherapy. Additionally, the period following treatment may allow for studies of minimal residual disease as well as the testing of potential maintenance therapies. Finally, this approach permits a detailed analysis of the critical few cancer stem cells that remain after induction therapy with the goal of identifying novel compounds capable of targeting these cells. Disclosures: No relevant conflicts of interest to declare.

Investigating the Expression of the MRD Marker CD58 on Leukaemia Initiating Cells in Childhood Acute Lymphoblastic Leukaemia

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1887-1887 ◽  
Author(s):  
Charlotte Victoria Cox ◽  
Paraskevi Diamanti ◽  
Allison Blair
Keyword(s):  
Acute Lymphoblastic Leukaemia ◽  
Lymphoblastic Leukaemia ◽  
Conflicts Of Interest ◽  
Residual Disease ◽  
Model Systems ◽  
Childhood All ◽  
Therapy Assessment ◽  
Childhood Acute Lymphoblastic Leukaemia

Abstract Abstract 1887 Further improvements in outcome for childhood acute lymphoblastic leukaemia (ALL) will require a better understanding of the underlying biology of this disease and the fundamental mechanisms of drug resistance. The discoveries that a few populations can initiate leukemia in mouse models and that new populations of leukaemia initiating cells (LIC) can be detected following an initial round of transplantation in these models raises important questions about the biology of the leukaemias. If several cell populations have LIC properties, what are the relationships of these populations to each other and which populations are most important to target with therapy? It will also be important to determine whether there is any correlation in the biological properties of LIC identified in the model systems with the response of the patients to therapy. Assessment of minimal residual disease (MRD) levels provides a sensitive measurement of early treatment response and permits detection of the in vivo selected drug resistant population. CD58 (leucocyte function-associated antigen 3; LFA-3) is a useful marker in MRD tracking of B cell precursor (BCP) ALL. CD58 is over expressed in these cases permitting discrimination of leukaemia blasts from normal B cells. In this study we investigated whether CD58 is expressed on LIC populations in childhood ALL. Expression of CD58 and CD34 was assessed in a cohort of 12 diagnostic samples with mixed prognoses and compared to levels detected in 11 normal bone marrow (NBM) samples. Levels of CD58 were significantly higher in the ALL cases (57.4±37.7%) than on NBM cells (21.1±12.2%; p=0.007). Likewise, the CD34+/CD58+ population was larger in ALL cases than in normal cells (22.2±34.7% and 0.25±0.25%, respectively; p=0.05). Cells from eight of the 12 patients, were sorted on the basis of expression or lack of expression of these markers and the functional ability of the sorted subpopulations was assessed in vitro and in vivo. On sorting, the majority of cells were CD34−/CD58− (43.7±39.2%), 20.7±30.7% were CD34−/CD58+, 19±14.3% were CD34+/CD58+ and the CD34+/CD58− population accounted for 16.6±35.3%. Unsorted cells and all 4 sorted populations were set up in long-term culture to assess proliferative capability and the in vivo propagating potential was assessed in NSG mice. All 4 sorted subpopulations proliferated over the 6 week period but the highest levels of expansion were observed in the cultures of CD34+/CD58+ (6–420 fold) and CD34+/CD58− (3–24 fold) cells. Cytogenetic analyses confirmed that leukaemia cells were maintained in the culture system. Results from the in vivo analyses on 5 cases to date indicate that all 4 subpopulations contain LIC. In these cases, higher levels of engraftment were observed with CD34+/CD58+ (up to 20%) and with CD34−/CD58− subpopulations (6.1-98%). Serial transplantation studies will determine whether there are differences in the repopulating and self-renewal abilities of these LIC. These findings suggest that using CD58 alone or in combination with CD34 would be insufficient to track disease progression in ALL. Incorporating additional markers that are commonly used in MRD panels will provide valuable information on LIC populations and facilitate development of improved disease monitoring. Disclosures: No relevant conflicts of interest to declare.

Effect of inducing HR deficiency using AVB500, a receptor tyrosine kinase AXL inhibitor, on response to olaparib in uterine serous cancer.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. e18108-e18108
Author(s):  
Michael Driscoll Toboni ◽  
Barbara Blachut ◽  
Mary M Mullen ◽  
Jo'an Tankou ◽  
Hollie M Noia ◽  
...  
Keyword(s):  
Treatment Period ◽  
Cell Lines ◽  
Day Treatment ◽  
Parp Inhibitor ◽  
Tumor Burden ◽  
Scid Mice ◽  
In Vivo Studies ◽  
Treatment Groups

e18108 Background: Evidence suggests DNA repair is a therapeutic target in endometrial cancer (EC). Given this, we determined whether combination therapy with AVB500, an AXL inhibitor, could improve response in a uterine serous cancer (USC) model. Methods: Two USC cell lines (ARK1 & ARK4) were treated with AVB500 (Aravive Biologics, Houston, TX) in combination with the poly ADP ribose polymerase (PARP) inhibitor, olaparib. Colony forming assays were assessed after 4 days of treatment with either AVB500 alone, olaparib alone or combination treatment (olaparib + AVB500); colonies were stained and absorbance was obtained to calculate relative cell viability using Graph Pad Prism. Baseline homologous recombination (HR) status was determined after radiating cells with 10Gy and identifying RAD51 foci by immunofluorescence (IF). Cell lines were considered to be HR proficient if over 30% of the cells expressed RAD51 ( > 5 foci per cell). IF was conducted using a Leica confocal microscope and foci were quantified using FociCounter. In vivo studies were performed using NOD-SCID mice injected with 1 x 107 ARK1 cells intraperitoneally followed by treatment q3 days for a 14 and 21 day treatment period. Treatment groups were vehicle control, AVB500 alone, olaparib alone and olaparib with AVB500. Results: The absorbance for olaparib + AVB500 was significantly less than the olaparib only group in two assays involving ARK1s (0.417nm vs 0.756nm, p = 0.001; 0.320nm vs 0.620nm, p = 0.008) as well as in ARK4s (0.186nm vs 0.641nm, p = 0.003). The HR assay indicated both cell lines were HR proficient. After baseline HR proficiency was established, the cell lines were pretreated with AVB500 prior to radiation. When compared to cells without treatment with AVB500, IF showed a decrease in RAD51 foci per cell in ARK1 (2.7 vs 7.3, p = 0.0003) and ARK4 (6.3 vs 13.0, p = 0.0054). The proportion of ARK1 cells expressing RAD51 decreased to 21%, indicating HR deficiency. Lastly, NOD-SCID mice receiving olaparib + AVB500 had less tumor weight than those treated with olaparib alone (0.008g vs 0.138g, p = 0.002) and AVB500 alone (0.008g vs 0.145g, p = 0.0006) in a 14 day and a 21 day treatment period (0.212g vs 0.586g, p = 0.027 and 0.212 vs 0.494g, p = 0.005, respectively). Conclusions: HR proficient USC cell lines treated in vitro and in vivo with the combination of AVB500 and olaparib demonstrate an improved response to olaparib or AVB500 alone with a greater decrease in tumor burden. AVB500 appears to induce HR deficiency. Additional therapeutic and mechanistic experiments are ongoing.

R-DHAP Is Effective in Fludarabine-Refractory CLL, Possibly Via Upregulation of Pro-Apoptotic P73.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3449-3449
Author(s):  
Sanne H. Tonino ◽  
Michel Van Gelder ◽  
Eric Eldering ◽  
Marinus H. J. van Oers ◽  
Arnon P. Kater
Keyword(s):  
Conflicts Of Interest ◽  
Residual Disease ◽  
Tumor Lysis Syndrome ◽  
Remission Induction ◽  
Refractory Disease ◽  
P53 Family ◽  
Bulky Disease ◽  
P53 Response

Abstract Abstract 3449 Poster Board III-337 Allogeneic stem cell transplantation (alloSCT) has the potential to cure high risk CLL patients including those with fludarabine-refractory disease with or without a deletion of 17p. Treatment-related mortality (TRM) of this procedure is significantly reduced by the use of reduced-intensity conditioning regimens (RIC). Risk factors for relapse after alloSCT are refractory or bulky disease. In an ongoing prospective Dutch/ Belgian HOVON trial we study the efficacy of remission-induction with the R-DHAP regimen (rituximab, cytarabine, cisplatinum and dexamethasone) prior to alloSCT in CLL patients with fludarabine refractory disease. During the initiation phase ten fludarabine refractory patients have been treated according to this protocol. The table shows their disease characteristics and response to R-DHAP. patient age FISH bulky disease1 preceding therapies No. of R-DHAP cycles Response 1 36 del 17p + Chl, CVP, FCR, Alem, R-CHOP 1 SD 2 65 normal - Chl, CVP, F 3 PR 3 71 normal + Chl, F, FC, FCR 1 PR 4 51 normal + Chl, F, FCR 4 SD 5 47 del 11q + FCR, Alem 3 PR 6 53 ND - Chl, F, Alem 3 PR 7 54 ND + Chl, F, FCR, R-CHOP 4 PR 8 65 ND + Chl, CVP, F, CHOP, Alem 3 PR 9 57 del 17p + R-CVP, R-CHOP 4 PR 10 50 del 11q + Chl, FCR, Alem 3 CR 1 lymph nodes > 5 cm Eight of ten patients had a response: one CR and seven PR, whereas two had SD. In the patient who achieved a CR, four-color flowcytometry confirmed the absence of minimal residual disease (MRD). One of the two patients with del 17p and both patients with del 11q had a response. Six of the eight patients with bulky disease responded. One patient developed tumor lysis syndrome. One patient, who had been pretreated with alemtuzumab (no 8), developed an opportunistic infection (Aspergillus pneumonia). Most responding patients subsequently underwent alloSCT. The observed response rate in these heavily pre-treated and chemo-refractory patients is remarkable. As chemorefractory disease is highly associated with a dysfunctional p53 response, we hypothesized that effects of this regimen could be independent of p53-function. Therefore the molecular basis of apoptosis induction was studied more in depth in one patient with del 17p and proven p53 dysfunction. Within weeks after the first cycle of R-DHAP peripheral blood lymphocyte counts decreased from 78 × 109/L to 3.3 × 109/L. Simultaneous analysis of RNA-expression levels of 30 apoptosis regulating genes by RT-MLPA in samples taken before and 24 and 48 hours after start of R-DHAP revealed significant upregulation of the pro-apoptotic BH3-only molecule Puma. Recent data indicate that Puma is not only a p53 response gene, but can also be regulated by the p53-family member p73. Indeed, detectable p73 protein levels were found already after 24 hours of treatment in vivo. At the functional level, we found that the pretreatment fludarabine resistance in vitro was abrogated after 48 hours of treatment in vivo. Our data indicate that the R-DHAP regimen has activity in fludarabine-refractory CLL patients, even in those with cytogenetic changes affecting the p53-response. Furthermore, in our preliminary in vitro studies we could show upregulation of the pro-apoptotic protein p73 and concomitant abrogation of fludarabine-resistance. This pathway should be further explored and may provide means to overcome drug-resistance in CLL. Disclosures No relevant conflicts of interest to declare.

Assessing CD97 and CD99 As Markers of Leukaemia Initiating Cells in Paediatric ALL

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1882-1882 ◽  
Author(s):  
Charlotte Victoria Cox ◽  
Paraskevi Diamanti ◽  
Allison Blair
Keyword(s):  
Functional Capacity ◽  
Conflicts Of Interest ◽  
Residual Disease ◽  
Survival Rates ◽  
In Vivo Studies ◽  
Nsg Mice ◽  
Early Treatment Response ◽  
Current Therapies

Abstract Abstract 1882 Overall survival rates in paediatric acute lymphoblastic leukaemia (ALL) have dramatically improved but around 20% do not respond to current therapies and subsequently relapse. Leukaemia initiating cells (LIC) are the topic of much investigation, as these cells can self-renew and may have the potential to cause relapse. It has been shown that multiple subpopulations of ALL cells have the ability to initiate the disease in immune deficient mouse models. Therefore, treatment should be targeted at all cells with this capacity, if the disease is to be eradicated. Minimal residual disease (MRD) detection is an invaluable tracking tool to assess early treatment response and recent studies have highlighted potential markers that may improve the sensitivity of MRD detection by flow cytometry. CD97 and CD99 are two markers which were over expressed in paediatric ALL. Incorporating these markers into investigations of LIC may allow discrimination of leukaemia cells from normal haemopoietic stem cells (HSC). In this study we evaluated the expression of CD34 in combination with CD97 in B cell precursor (BCP) ALL cases and CD99 in T-ALL cases and subsequently assessed the functional capacity of the sorted subpopulations in vitro and in vivo. Ten ALL samples (6 B-ALL & 4 T-ALL) with a median age 7 years (range 2–15 years) were studied. One B-ALL case and 3 T-ALL cases were considered high risk by molecular assessment of MRD at day 28 of treatment. Flow cytometric analyses of the ALL samples and 8 normal haemopoietic cell samples demonstrated that both CD97 and CD99 were over expressed in ALL patients (78.9±14.8% & 76.4±32.8%, respectively) when compared to normal haemopoietic cells (14.1±25.4%; p=0.001, 47.1±10%; p=0.03, respectively). Cells were sorted for expression/lack of expression of these markers and proliferation of the sorted cells was assessed in suspension culture over a 6 week period. In the B-ALL patients the CD34+/CD97+ subpopulation represented the bulk of leukaemia cells (65.2±32.1%), the CD34−/CD97+ the smallest fraction (3.3±2.4%) with the CD34+/CD97− and CD34−/CD97− subpopulations representing 21.1±31.5% and 10.5±5.8% of cells, respectively. When the functional capacity of these subpopulations was assessed in vitro greatest expansion was observed in cells derived from CD34+/CD97− subpopulation (2–173 fold) from 9.4×103 at initiation up to 1.5×106 cells at week 6. Expansion was also observed, to a lesser extent in the CD34−/CD97− subpopulation (3.4–28 fold) from 8×103 up to 1.4×106 cells. No expansion was observed in cultures of CD34+/CD97+ and CD34−/CD97− subpopulations but cells were maintained throughout the culture period. These sorted subpopulations were also inoculated into NOD/LtSz-SCID IL-2Rγc null (NSG) mice to evaluate repopulating capacity. To date, engraftment has been achieved with 3 subpopulations; CD34+/CD97+ (3–28.8% CD45+), CD34+/CD97− (0.5–25.5% CD45+) and CD34−/CD97+ (23.8% CD45+) cells. When the functional capacity of T-ALL cases was assessed the CD34+/CD99+ subpopulation represented the bulk of cells at sorting (51.87±47.2%), the CD34+/CD99- subpopulation was the smallest (0.9±0.8%) and the CD34−/CD99+ and CD34−/CD99− subpopulations represented 32.1±38.9% and 27.2±33.4% of cells, respectively. Greatest expansion was observed in cultures of CD34+/CD99- cells (4.6–1798 fold) from 7.5×103 up to 2.6×106 cells at week 6. The other 3 subpopulations expanded to a lesser extent (1.3–216 fold) from 5×103 up to 1.8×106 cells. When the functional capacity of these cells was assessed in NSG mice, engraftment was achieved in all subpopulations; CD34+/CD99+ (87–90.5% CD45+), CD34+/CD99− (1.5–84.9% CD45+), CD34−/CD99+ (31.3–98.6% CD45+) and CD34−/CD99− (3–92.9% CD45+). In some cases, cells recovered from BM of NSG inoculated with CD99− cells had high expression of CD99, typical of the patient samples at diagnosis, indicating that the inoculated CD99− cells had differentiated in vivo. Studies are ongoing to assess the self-renewal capacity of these subpopulations by serial transplantation. The findings to date indicate that targeting CD97 and CD99, either alone or in combination with CD34 would not eliminate all cells with the capacity to initiate and maintain B-ALL and T-ALL, respectively. Further developments in therapy may require targeting leukaemogenic pathways, rather than only cell surface markers to improve survival outcome in paediatric ALL. Disclosures: No relevant conflicts of interest to declare.

Nanomagnetic liposome-encapsulated parthenolide and indocyanine green for targeting and chemo-photothermal antitumor therapy

Nanomedicine ◽  
2020 ◽  
Vol 15 (9) ◽  
pp. 871-890 ◽  
Author(s):  
Wenbin Gao ◽  
Lei Li ◽  
Xuwu Zhang ◽  
Liyao Luo ◽  
Yuchu He ◽  
...  
Keyword(s):  
Indocyanine Green ◽  
Synergistic Effects ◽  
Day Treatment ◽  
Antitumor Efficacy ◽  
Magnetic Targeting ◽  
Antitumor Therapy ◽  
Antitumor Effects ◽  
Targeting Therapy

Aim: To synthesize a drug-delivery system with chemo-photothermal function and magnetic targeting, to validate its antitumor effect. Materials & methods: Parthenolide (PTL), employing chemotherapy and indocyanine green (ICG) providing phototherapy, were encased separately in the lipid and aqueous phases of liposomes (Lips). The Fe3O4 nanoparticles (MNPs), endowing magnetic targeting, were modified on the surface of Lips. The antitumor effects were investigated in vitro and in vivo. Results: ICG-PTL-Lips@MNPs showed outstanding synergistic antitumor efficacy in vitro and in vivo. Especially, after 14-day treatment, the tumor volumes decreased significantly and the biotoxicity was very low. Conclusion: The designed ICG-PTL-Lips@MNPs possess synergistic effects of chemotherapy, photothermal and targeting therapy, which are expected to provide an alternative way to further improve antitumor efficacy.

scholarly journals Anti-cancer properties of cannabidiol and Δ9-tetrahydrocannabinol and potential synergistic effects with gemcitabine, cisplatin and other cannabinoids in bladder cancer

2021 ◽  
Author(s):  
Andrea M Tomko ◽  
Erin G Whynot ◽  
Denis J Dupre
Keyword(s):  
Bladder Cancer ◽  
Cell Viability ◽  
Synergistic Effects ◽  
Transitional Cell ◽  
Chemotherapeutic Agents ◽  
Anti Cancer ◽  
Concentration Curves ◽  
Apoptotic Cascade

Introduction: With the legalization of cannabis in multiple jurisdictions throughout the world, a larger proportion of the population consumes cannabis. Several studies have demonstrated anti-tumor effects of components present in cannabis in different models. Unfortunately, little is known about the potential anti-tumoral effects of cannabinoids in bladder cancer, and how cannabinoids could potentially synergize with chemotherapeutic agents. Our study aims to identify whether a combination of cannabinoids, like cannabidiol and Δ9-tetrahydrocannabinol with agents commonly used to treat bladder cancer, such as gemcitabine and cisplatin, is able to produce desirable synergistic effects. We also evaluated whether co-treatment of different cannabinoids also generated synergistic effects. Materials and Methods: We generated concentration curves with different drugs to identify the range at which they could exert anti-tumor effects. We also evaluated the activation of the apoptotic cascade and whether cannabinoids have the ability to reduce invasion. Results: Cannabidiol, Δ9-tetrahydrocannabinol and other cannabinoids reduce cell viability of bladder cancer cell lines, and their combination with gemcitabine or cisplatin may induce differential responses: from antagonistic to additive and synergistic effects, depending on the concentrations used. Cannabidiol and Δ9-tetrahydrocannabinol were also shown to induce caspase 3 cleavage and reduce invasion in a Matrigel assay. Cannabidiol and Δ9-tetrahydrocannabinol also display synergistic properties with other cannabinoids like cannabichromene or cannabivarin. Discussion: Our results indicate that cannabinoids can reduce human bladder transitional cell carcinoma cell viability, and that they can potentially exert synergistic effects when combined with other agents. Our in vitro results will form the basis for future studies in vivo and in clinical trials for the development of new therapies that could be beneficial for the treatment of bladder cancer in the future.

Growth Inhibition and Synergistic Induction of Apoptosis By Synthetic Mir-125b-5p Mimics and Myc-Targeting Agents in Human Myeloma Cell Lines

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3019-3019
Author(s):  
Lavinia Biamonte ◽  
Cinzia Federico ◽  
Eugenio Morelli ◽  
Emanuela Leone ◽  
Maria Eugenia Gallo Cantafio ◽  
...  
Keyword(s):  
Cell Lines ◽  
Conflicts Of Interest ◽  
Therapeutic Potential ◽  
Apoptotic Cell ◽  
Synergistic Effects ◽  
Annexin V ◽  
Transcriptional Level ◽  
Suppressor Activity

Abstract MicroRNAs (miRNAs), short non-coding RNAs which tune gene expression at post-transcriptional level, are recently emerging as key players in pathogenesis, progression and drug-resistance of multiple myeloma (MM). In this disease, they can act either with tumor-promoting or tumor-suppressing functions, depending on the nature of target mRNAs. Nowadays, effective strategies are available both to replace or to inhibit the expression of deregulated miRNAs, thus prompting the design of miRNA-based therapeutic strategies. We have recently demonstrated that miR-125b has tumor suppressor activity in MM and that enforced expression of synthetic miR-125b-5p mimics induces significant anti-MM activity in vitro and in vivo by targeting cell addiction to IRF4/cMyc pro-survival signaling. Moreover, we uncovered a functional feedback loop between cMyc and miR-125b in MM cells, whereas cMyc directly suppresses miR-125b transcription which, in turn, negatively regulates cMyc expression by targeting IRF4 mRNA. In the present study, we investigated the therapeutic potential of synthetic miR-125b-5p mimics combined with cMyc targeting agents, including the 10058-F4 small molecule inhibitor of cMyc-Max heterodimerization and the BET-bromodomain inhibitor JQ1, which is reported to inhibit cMyc transcription. At this aim, 3 MM cell lines (NCI-H929, SK-MM-1 and RPMI-8226) transfected with either miR-125b-5p mimics or scrambled oligonucleotides (miR-NC) were exposed to 10058-F4 (ranging from 10 to 100 μM) or JQ1 (ranging from 0,1 to 2μM) or DMSO. Effects on cell proliferation were then evaluated by CCK-8 assay at 24h, 48h and 72h time points, while the occurrence of apoptotic cell death was assessed by Annexin V flow-cytometry assay. Importantly, we found that enforced expression of miR-125b-5p mimics significantly and synergistically (synergistic index, SI >1) increases growth-inhibitory and pro-apoptotic activities of both 10058-F4 and JQ1. Similar results were observed in SK-MM-1 cells co-transfected with miR-125b-5p and cMyc siRNAs, while cMyc-defective U266 cells were not sensitized to either 10058-F4 nor JQ1 upon transfection with miR-125b-5p mimics. Furthermore, combinatorial treatments with JQ1 and miR-125b-5p mimics resulted in a stronger downregulation of cMyc protein, as compared to single molecules alone. Indeed, these results confirmed that impairment of cMyc activity/expression mediates the anti-MM synergistic effects between 10058-F4 or JQ1 and overexpression of miR-125b-5p by synthetic mimics. In conclusion, our data demonstrate a cMyc-mediated synergistic anti-MM activity of synthetic miR-125b-5p mimics with 10058-F4 or JQ1 cMyc targeting agents, providing the rationale for a more advanced preclinical investigations for the design of early clinical trials. Disclosures No relevant conflicts of interest to declare.

scholarly journals Polymeric Nanoparticle Delivery of Combination Therapy with Synergistic Effects in Ovarian Cancer

Nanomaterials ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 1048
Author(s):  
Shani L. Levit ◽  
Christina Tang
Keyword(s):  
Ovarian Cancer ◽  
Drug Resistance ◽  
Treatment Efficacy ◽  
Synergistic Effects ◽  
Chemotherapeutic Agents ◽  
Drug Combinations ◽  
Dosing Schedule ◽  
Acquired Drug Resistance

Treatment of ovarian cancer is challenging due to late stage diagnosis, acquired drug resistance mechanisms, and systemic toxicity of chemotherapeutic agents. Combination chemotherapy has the potential to enhance treatment efficacy by activation of multiple downstream pathways to overcome drug resistance and reducing required dosages. Sequence of delivery and the dosing schedule can further enhance treatment efficacy. Formulation of drug combinations into nanoparticles can further enhance treatment efficacy. Due to their versatility, polymer-based nanoparticles are an especially promising tool for clinical translation of combination therapies with tunable dosing schedules. We review polymer nanoparticle (e.g., micelles, dendrimers, and lipid nanoparticles) carriers of drug combinations formulated to treat ovarian cancer. In particular, the focus on this review is combinations of platinum and taxane agents (commonly used first line treatments for ovarian cancer) combined with other small molecule therapeutic agents. In vitro and in vivo drug potency are discussed with a focus on quantifiable synergistic effects. The effect of drug sequence and dosing schedule is examined. Computational approaches as a tool to predict synergistic drug combinations and dosing schedules as a tool for future nanoparticle design are also briefly discussed.

Activation Of Natural Killer Cells By Soypeptide Lunasin and Cytokine: Implication In Cancer Immunotherapy For Lymphoma

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 1042-1042
Author(s):  
Chun-Yu Tung ◽  
Sharifah Kyazike ◽  
David Lewis ◽  
Ling Han ◽  
Alexander Kolb ◽  
...  
Keyword(s):  
Nk Cells ◽  
Natural Killer ◽  
Conflicts Of Interest ◽  
Synergistic Effects ◽  
Underlying Mechanism ◽  
Tumor Models ◽  
Tumoricidal Activity ◽  
Ifng Production

Abstract Introduction Immunostimulatory cytokines can enhance anti-tumor immunity and are part of the therapeutic armamentarium for cancer treatment. We have previously reported that chemotherapy-treated lymphoma patients have acquired deficiency of Signal Transducer and Activator of Transcription 4 (STAT4), which results in defective IFNg production during clinical immunotherapy. With the goal of further improvement in cytokine-based immunotherapy, we examined the effects of a soybean peptide called lunasin that exhibits immunostimulatory effects on natural killer (NK) cells. Experimental Design PBMCs of healthy donors and chemotherapy-treated lymphoma patients were stimulated with or without lunasin in the presence of IL-12 or IL-2. NK activation was evaluated, and its tumoricidal activity was assessed using in vitro and in vivo tumor models. Chromatin immunoprecipitation (ChIP) assay was performed to evaluate the histone modification of gene loci that are regulated by lunasin and cytokine. Results Adding lunasin to IL-12- or IL-2-cultuted NK cells demonstrated synergistic effects in the induction of IFNG and genes involved in cytotoxicity. The expression level of CD16 and granzyme B was increased in CD56-bright population of NK cells following stimulation with lunasin and cytokine. The combination of lunasin and cytokines (IL-12 plus IL-2) was capable of restoring IFNg production by NK cells from post-transplant lymphoma patients. In addition, NK cells stimulated with lunasin plus cytokines had higher tumoricidal activity than those stimulated with cytokines alone using in vitro and in vivo tumor models. Moreover, lunasin augmented antibody-dependent cellular cytotoxicity (ADCC) of NK cells against anti-CD20 coated human B-lymphoma cell line. The underlying mechanism responsible for the effects of lunasin on NK cells is likely due to epigenetic modulation on target gene loci. Conclusion We have discovered a novel use of lunasin that exerts synergistic effects together with IL-12 or IL-2. This synergism leads to stronger NK-mediated anti-tumor activity, suggesting the potential clinical use of lunasin to augment the therapeutic responses in cytokine-based immunotherapy. Disclosures: No relevant conflicts of interest to declare.

Bifunctional Immunoactive siRNAs as an Approach to Personalized AML Therapy

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 5036-5036
Author(s):  
Hanna Meinl ◽  
Marcus Zeitlhöfler ◽  
Samet Kocabey ◽  
Tim Liedl ◽  
Wolfgang Hiddemann ◽  
...  
Keyword(s):  
Target Genes ◽  
Conflicts Of Interest ◽  
Residual Disease ◽  
Genetic Alterations ◽  
Chemotherapeutic Agents ◽  
Leukemic Cells ◽  
In Vivo Studies ◽  
Specific Gene ◽  
Therapeutic Concepts

Abstract The prognosis of acute myeloid leukemia (AML) is poor due to frequent relapse after initial remission. The development of new approaches to postremission therapy for elimination of minimal residual disease remains a major scientific and clinical challenge. We strive to combine two different innovative therapeutic concepts to develop a new specific and personalized treatment for AML. siRNAs are used to knock down either a gene that drives leukemogenesis due to genetic alterations in specific cases of AML (e.g., FLT3, NPM1) or a gene that is essential for the survival of the leukemic cells (e.g., BRD4, MCL1, PLK1). By adding a triphosphate modification to the 5’ end, the siRNA molecules additionally become ligands for the cytosolic pattern recognition receptor RIG-I (retinoic acid inducible gene I). Its activation mimics viral infection and leads to the production of inflammatory cytokines and induction of apoptosis in the target cell. We expect these bifunctional molecules to result in a decrease of viable AML cells and in the induction of an immune response similar to an active immunization. This concept was successfully tested in vitro for several target genes in AML cell lines. We could demonstrate that the specific gene knockdown leads to inhibited proliferation, increased apoptosis and higher sensitivity to chemotherapeutic agents. Activation of RIG-I by triphosphate-modified RNA additionally stimulated an inflammatory response by the leukemic cells and increased the apoptosis rate. A major hurdle for all siRNA-based anti-cancer strategies is the specific delivery of the RNA into tumor cells. In vivo liposomal transfection of siRNA molecules has been used in various tumor models, but generally results in ineffective and unspecific delivery. We are testing DNA-based nanoparticles coupled with molecules that target receptors specific for or overexpressed on AML cells. By coupling bifunctional siRNA molecules to these nanoparticles, they should be efficiently and selectively transported into the cytosol of AML cells. Proof-of-concept in vivo studies in AML mouse models are in preparation. The long-term goal of this project is the development of a set of bifunctional siRNA molecules for the individualized treatment of AML. Disclosures: No relevant conflicts of interest to declare.

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