19 abril 2018

PharmaMar en el Congreso de la AACR . 16 al 18 de Abril en Chicago . PM184 ( PLOCABULIN ) y Aplidin .

Miercoles 18 Abril 2018 :

Exploratory testing of PM060184 (PLOCABULIN ) .

Compound in high-grade serous ovarian carcinoma cell lines .

 April 18, 2018 . Section 39
V. Heredia-Soto1, A. Redondo1, A. Gallego1, M. Miguel-Martín1, R. Crespo1, A. Hernández1, D. Hardisson1, J. Feliu1, C. Galmarini2M. Mendiola1;1La Paz Hosp., IdiPAZ, Madrid, Spain, 2Pharmamar, Madrid, Spain
 V. Heredia-Soto: None. A. Redondo: ; Pharmamar. A. Gallego: None. M. Miguel-Martín: None. R. Crespo: None. A. Hernández:None. D. Hardisson: None. J. Feliu: None. C. Galmarini: ; Pharmamar. M. Mendiola: ; Pharmamar.

Purpose of the Study: Marine sponges have developed mechanisms to protect themselves from a hostile marine microenvironment. One of these mechanisms is the use of biologically active metabolites, explored nowadays for their anticancer properties. PM060184 is one of these compounds, isolated from the Madagascan sponge Lithoplocamia lithistoides. This polyketide is currently under evaluation on clinical trials, and its antitumor activity was previously reported in the preclinical setting in a panel of cell lines and subcutaneous tumor xenografts of different origin. The purpose of the study is to explore the effect of PM060184 in a panel of ovarian high-grade serous (HGS) carcinoma cell lines with different sensitivity to cisplatin and paclitaxel.
Experimental Procedures: Cell growth inhibition was analyzed by exposure of increasing concentrations of PM060184 in 96 multiwell plates for 72h, by subsequent confluence evaluation and sulphorhodamine (SRB) staining. Cell cycle experiments were done by propidium iodide (PI) staining after treatment of cell lines at their IC50 value for 72h. Three independent experiments with 6 replicates per condition were performed for both approaches. Celigo Image Cytometer platform was employed for phase contrast and viable cells discrimination with a triple staining (Hoechst, PI and calcein AM), as well as for cell cycle analyses.
Results: We have focused on the effect of this drug on a panel of HGS ovarian carcinoma cell lines, previously characterized by their response to cisplatin and paclitaxel. We have observed antiproliferative activity in a concentration-dependent manner, with IC50 values at subnanomolar concentrations in all the cell lines tested. This effect was observed in platinum-sensitive and -resistant cell lines and all of them were also more sensitive to PM060184 than paclitaxel, another tubulin-binding agent usually used for the treatment of ovarian cancer. We have evaluated cell cycle and apoptosis at the IC50values for each cell line. We have seen that this agent disrupts the cell cycle at different phases, as DNA synthesis and mitosis, depending on the cell line. Additionally, an increase in apoptotic cell death is detected as a sub G1 peak by flow cytometry in most of the cell lines tested.
Conclusions: PM060184 is a new tubulin-binding agent with a potent antitumor activity in a panel of HGS ovarian cancer cell lines, with different ranges of sensitivity to cisplatin and paclitaxel. Additionally, this effect is more potent than that exerted by other tubulin-binding compounds, such as paclitaxel. However, the underlying mechanism of PM060184 action on ovarian HGS cell lines will need to be further elucidated.


Lunes 16 Abril 2018 :

Session PO.ET01.06 - New Targets 2

APLIDIN (Plitidepsin).

eEF1A2 interacts with and inhibits PKR to enhance cancer cell survival .

 April 16, 2018, 1:00 PM - 5:00 PM Section 40

J. F. Martínez-Leal, A. Losada, M. Muñoz, M. Martinez-Diez, J. Dominguez, C. M. Galmarini; PharmaMar S.A., Madrid, Spain.

  J.F. Martínez-Leal: ; Pharmamar. A. Losada: ; Pharmamar. M. Muñoz: ; Pharmamar. M. Martinez-Diez: ; Pharmamar. J. Dominguez: ; Pharmamar. C.M. Galmarini: ; Pharmamar.

Human translation elongation factor 1α2, encoded by the eEF1A2 gene, is a pro-oncogenic protein absent from the majority of body tissues (with exception of brain, heart and skeletal muscle1), but expressed in many cancers1-3, where it provides tumor cells with improved fitness and survival. Though its canonical function is delivering aminoacyl-tRNAs to the ribosome, other “moonlighting” functions such as enhancing sphingosine kinase4,5 or antioxidant (most probably through peroxiredoxin-1 stimulation) activities6have been described for the elongation factor. Recently, we have reported that eEF1A2 is the target for plitidepsin, a marine-derived cyclic depsipeptide currently under development for the treatment of relapsed or refractory multiple myeloma patients7. We have also confirmed that eEF1A2 interacted with previously described partners as PRDX1 and SPHK and enhanced their pro-survival activities8. Here we investigated the role of new “moonlighting functions” of eEF1A2 in the maintenance of the tumor phenotype and survival of cancer cells. Through co-immunoprecipitation and HPLC/MS we have uncovered the interaction between eEF1A2 and dsRNA-activated protein kinase (PKR, EIF2AK2). We have analyzed the kinase activity of PKR in the presence of eEF1A2, demonstrating that PKR activity is inhibited when complexed with eEF1A2. This complex is disrupted after plitidepsin binding to eEF1A2, rendering PKR active. Once activated, the kinase triggers a MAPK cascade and the NF-κB signaling pathway, leading to the activation of the extrinsic apoptotic pathway and the death of the tumor cell. Taken together, these results show that the fitness boost that the moonlighting functions of eEF1A2 provide to cancer cells, which are important for their growth and survival, constitutes an Achilles heel that can be purposely exploited in anticancer therapy. 


Martes 17 Abril 2018 :

Plocabulin ( PLO ) , a tubulin inhibitor, presents antitumor activity in patient-derived xenograft (PDX) models of gastrointestinal stromal tumor (GIST) .

 April 17, 2018, 8AM - 12:00 PM

A. Wozniak1, Y. Wang1, J. Wellens1, Y. K. Gebreyohannes1, M. Guillén2, C. M. Galmarini2, P. M. Avilés2, M. Debiec-Rychter1, R. Sciot1, P. Schöffski11KU Leuven and Univ. Hosp. Leuven, Leuven, Belgium, 2PharmaMar, Madrid, Spain
 A. Wozniak: None. Y. Wang: None. J. Wellens:None. Y.K. Gebreyohannes: None. M. Guillén: ; PharmaMar. C.M. Galmarini: ; PharmaMar. P.M. Avilés: ; PharmaMar. M. Debiec-Rychter:None. R. Sciot: None. P. Schöffski: ; travel grants All institutional funding; PharmaMar.
Introduction: Advanced GIST is commonly treated with tyrosine kinase inhibitors (TKI) [e.g. imatinib (IMA)]. With time the vast majority of patients develops TKI-resistance. GIST is generally believed to be resistant to chemotherapy with cytotoxic agents. The aim of our study was to test plocabulin (PLO; PM060184, PharmaMar), a potent cytotoxic tubulin-dynamics modifier, in two PDX models of GIST, characterized by different sensitivity to IMA.

Experimental set-up: NMRI nu/nu mice (n=34) were transplanted bilaterally with human xenografts UZLX-GIST3sens (KIT: exon 11 p.W557_V559delinsF; IMA-sensitive) or -GIST9res(KIT: exon 11+17: p.P577del;W557LfsX5;D820G; IMA-resistant). Xenografted animals were randomly assigned to three treatment groups: control [vehicle, 5ml/kg/QW intravenously (i.v.)], IMA (50mg/kg/BID orally) and PLO (16mg/kg/QW, i.v.). Treatment lasted 22 days and the antitumor activity was assessed by tumor volume measurement, histopathology and KIT signaling pathway by Western blotting. Histological response (HR) was evaluated as previously described by Antonescu et al. 2005. Mann Whitney U test was used for statistical analysis with p <0.05 considered as significant.

Results: PLO treatment resulted in a reduction of tumor volume to 59% in GIST3sens and to 70% of the baseline volume in the GIST9res model. Good HR (grade 3, 4) was observed in 70% (GIST3sens) and 50% (GIST9res) of tumors. HR obtained with PLO was mainly characterized by necrosis, while IMA produced mainly myxoid degeneration in the sensitive model. In addition, in GIST3sens PLO decreased the microvessel area and increased apoptosis as assessed by immunohistochemistry, which was not observed in GIST9res. In the latter model PLO showed better activity than IMA in terms of tumor volume reduction (59% vs. 146%, p<0.01) and HR (grade 3, 4 in 50% vs. 0%). KIT signaling was not affected by PLO. The experimental drug was well tolerated throughout the experiment at the dose administered.

Conclusions: PLO is the first anti-tubulin agent showing antitumor activity in GIST PDX, both in models sensitive or resistant to IMA. The drug causes cytotoxicity in GIST, mainly through necrosis, without affecting KIT signaling. Due to the different modes of action of PLO and established TKI our work provides a scientific rationale to combine PLO and IMA to overcome resistance to small molecule TKI.