Potent Antitumor Effects Research Articles

A Cancer-Specific Monoclonal Antibody against Podocalyxin Exerted Antitumor Activities in Pancreatic Cancer Xenografts.

Podocalyxin (PODXL) overexpression is associated with poor clinical outcomes in various tumors. PODXL is involved in tumor malignant progression through the promotion of invasiveness and metastasis. Therefore, PODXL is considered a promising target of monoclonal antibody (mAb)-based therapy. However, PODXL also plays an essential role in normal cells, such as vascular and lymphatic endothelial cells. Therefore, cancer specificity or selectivity is required to reduce adverse effects on normal cells. Here, we developed an anti-PODXL cancer-specific mAb (CasMab), PcMab-6 (IgG1, kappa), by immunizing mice with a soluble PODXL ectodomain derived from a glioblastoma LN229 cell. PcMab-6 reacted with the PODXL-positive LN229 cells but not with PODXL-knockout LN229 cells in flow cytometry. Importantly, PcMab-6 recognized pancreatic ductal adenocarcinoma (PDAC) cell lines (MIA PaCa-2, Capan-2, and PK-45H) but did not react with normal lymphatic endothelial cells (LECs). In contrast, one of the non-CasMabs, PcMab-47, showed high reactivity to both the PDAC cell lines and LECs. Next, we engineered PcMab-6 into a mouse IgG2a-type (PcMab-6-mG2a) and a humanized IgG1-type (humPcMab-6) mAb and further produced the core fucose-deficient types (PcMab-6-mG2a-f and humPcMab-6-f, respectively) to potentiate the antibody-dependent cellular cytotoxicity (ADCC). Both PcMab-6-mG2a-f and humPcMab-6-f exerted ADCC and complement-dependent cellular cytotoxicity in the presence of effector cells and complements, respectively. In the PDAC xenograft model, both PcMab-6-mG2a-f and humPcMab-6-f exhibited potent antitumor effects. These results indicated that humPcMab-6-f could apply to antibody-based therapy against PODXL-expressing pancreatic cancers.

Pyrotinib shows a potent antitumor effect on HER2-positive esophageal cancer cells by promoting HER2 proteasomal degradation.

Pan-HER TKIs (pyrotinib, lapatinib) are potent HER2 inhibitors, however, their anti-tumor efficacy on esophageal cancer remains to be elucidated. Using two HER2-positive esophageal cancer cell lines, we observed that both pyrotinib and lapatinib could significantly suppress the activation of HER2 and its downstream signaling. However, pyrotinib showed a potent inhibitory effect at 0.1 µM treatment relative to 1 µM of lapatinib. Moreover, treatment with pyrotinibm, but not lapatinib, markedly reduced the protein level of HER2 through enhancing HER2 ubiquitination level and proteasomal degradation. In vitro and in vivo experiments further revealed that pyrotinib effectively suppresses cancer cell invasion and migration, as well as the growth of tumors in nude mice. Overall, our results suggest that pyrotinib is a superior TKI over lapatinib in inhibiting esophageal cancer cell proliferation and tumorigenic potential, and can be chosen as a neo-adjuvant for esophageal cancer treatment.

Exploring the Impact of Onobrychis cornuta and Veratrum lobelianum Extracts on C. elegans: Implications for MAPK Modulation, Germline Development, and Antitumor Properties.

In an era of increasing interest in the potential health benefits of medicinal foods, the need to assess their safety and potential toxicity remains a critical concern. While these natural remedies have garnered substantial attention for their therapeutic potential, a comprehensive understanding of their effects on living organisms is essential. We examined 316 herbal extracts to determine their potential nematocidal attributes in Caenorhabditis elegans. Approximately 16% of these extracts exhibited the capacity to induce diminished survival rates and larval arrest, establishing a correlation between larval arrest and overall worm viability. Certain extracts led to an unexpected increase in male nematodes, accompanied by a discernible reduction in DAPI-stained bivalent structures and perturbed meiotic advancement, thereby disrupting the conventional developmental processes. Notably, Onobrychis cornuta and Veratrum lobelianum extracts activated a DNA damage checkpoint response via the ATM/ATR and CHK-1 pathways, thus hindering germline development. Our LC-MS analysis revealed jervine in V. lobelianum and nine antitumor compounds in O. cornuta. Interestingly, linoleic acid replicated phenotypes induced by O. cornuta exposure, including an increased level of pCHK-1 foci, apoptosis, and the MAPK pathway. Mutants in the MAPK pathway mitigated the decline in worm survival, underscoring its importance in promoting worm viability. This study reveals complex interactions between herbal extracts and C. elegans processes, shedding light on potential antitumor effects and mechanisms. The findings provide insights into the complex landscape of herbal medicine's impact on a model organism, offering implications for broader applications.

Augmented Concentration of Isopentyl-Deoxynyboquinone in Tumors Selectively Kills NAD(P)H Quinone Oxidoreductase 1-Positive Cancer Cells through Programmed Necrotic and Apoptotic Mechanisms.

Lung and breast cancers rank as two of the most common and lethal tumors, accounting for a substantial number of cancer-related deaths worldwide. While the past two decades have witnessed promising progress in tumor therapy, developing targeted tumor therapies continues to pose a significant challenge. NAD(P)H quinone oxidoreductase 1 (NQO1), a two-electron reductase, has been reported as a promising therapeutic target across various solid tumors. β-Lapachone (β-Lap) and deoxynyboquinone (DNQ) are two NQO1 bioactivatable drugs that have demonstrated potent antitumor effects. However, their curative efficacy has been constrained by adverse effects and moderate lethality. To enhance the curative potential of NQO1 bioactivatable drugs, we developed a novel DNQ derivative termed isopentyl-deoxynyboquinone (IP-DNQ). Our study revealed that IP-DNQ treatment significantly increased reactive oxygen species generation, leading to double-strand break (DSB) formation, PARP1 hyperactivation, and catastrophic energy loss. Notably, we discovered that this novel drug induced both apoptosis and programmed necrosis events, which makes it entirely distinct from other NQO1 bioactivatable drugs. Furthermore, IP-DNQ monotherapy demonstrated significant antitumor efficacy and extended mice survival in A549 orthotopic xenograft models. Lastly, we identified that in mice IP-DNQ levels were significantly elevated in the plasma and tumor compared with IB-DNQ levels. This study provides novel preclinical evidence supporting IP-DNQ efficacy in NQO1+ NSCLC and breast cancer cells.

Drug repositioning in thyroid cancer treatment: the intriguing case of anti-diabetic drugs.

Cancer represents the main cause of death worldwide. Thyroid cancer (TC) shows an overall good rate of survival, however there is a percentage of patients that do not respond or are refractory to common therapies. Thus new therapeutics strategies are required. In the past decade, drug repositioning become very important in the field of cancer therapy. This approach shows several advantages including the saving of: i) time, ii) costs, iii) de novo studies regarding the safety (just characterized) of a drug. Regarding TC, few studies considered the potential repositioning of drugs. On the other hand, certain anti-diabetic drugs, were the focus of interesting studies on TC therapy, in view of the fact that they exhibited potential anti-tumor effects. Among these anti-diabetic compounds, not all were judjed as appropriate for repositioning, in view of well documented side effects. However, just to give few examples biguanides, DPP-4-inhibitors and Thiazolidinediones were found to exert strong anti-cancer effects in TC. Indeed, their effects spaced from induction of citotoxicity and inhibition of metastatic spread, to induction of de-differentiation of TC cells and modulation of TC microenvironment. Thus, the multifacial anti-cancer effect of these compounds would make the basis also for combinatory strategies. The present review is aimed at discuss data from studies regarding the anti-cancer effects of several anti-diabetic drugs recently showed in TC in view of their potential repositioning. Specific examples of anti-diabetic repositionable drugs for TC treatment will also be provided.

Antitumor and Phytochemical Properties of Ferula assa-foetida L. Oleo-Gum-Resin against HT-29 Colorectal Cancer Cells In Vitro and in a Xenograft Mouse Model.

Colorectal cancer (CRC) is one of the most frequently occurring tumors. Ferula assa-foetida oleo-gum-resin (OGR) extract is a traditional cooking spice known for its broad spectrum of biological activities such as antifungal, antiparasitic, and anti-inflammatory activities. This study evaluated the antitumor effect of OGR extract against HT-29 colorectal cancer cells. The OGR chemical composition was analyzed using LC-ESI-MS/MS; MTT, clonogenic assays, and a xenograft model were used to measure cytotoxicity, while apoptotic proteins were detected using Western blotting. Phytochemical analysis revealed that the extract was a rich source of isoflavones, xanthones, and other derivatives. In a dose-dependent manner, the OGR extract significantly inhibited colony formation ability and HT-29 cell growth (IC50 was 3.60 ± 0.02 and 10.5 ± 0.1 mg/mL, respectively). On the other hand, the OGR extract significantly induced apoptosis and increased the expression of some pro-death proteins involved in cellular apoptosis including PUMA, BIM, BIK, and BAK. Moreover, in a subcutaneous HT-29 xenograft model, the tumor volume and burden decreased after treatment with the OGR extract (550 ± 32 mm3 and 16.3 ± 3.6, respectively) This study demonstrated that Ferula assa-foetida OGR ethanolic extract has potential antitumor effects against HT-29 CRC cell lines by reducing cell viability and the function of apoptosis. More studies are needed to reveal the underlying mechanisms related to cytotoxicity and apoptosis induction.

The Emerging Roles of the Metabolic Regulator G6PD in Human Cancers.

Metabolic reprogramming, especially reprogrammed glucose metabolism, is a well-known cancer hallmark related to various characteristics of tumor cells, including proliferation, survival, metastasis, and drug resistance. Glucose-6-phosphate dehydrogenase (G6PD) is the first and rate-limiting enzyme of the pentose phosphate pathway (PPP), a branch of glycolysis, that converts glucose-6-phosphate (G6P) into 6-phosphogluconolactone (6PGL). Furthermore, PPP produces ribose-5-phosphate (R5P), which provides sugar-phosphate backbones for nucleotide synthesis as well as nicotinamide adenine dinucleotide phosphate (NADPH), an important cellular reductant. Several studies have shown enhanced G6PD expression and PPP flux in various tumor cells, as well as their correlation with tumor progression through cancer hallmark regulation, especially reprogramming cellular metabolism, sustaining proliferative signaling, resisting cell death, and activating invasion and metastasis. Inhibiting G6PD could suppress tumor cell proliferation, promote cell death, reverse chemoresistance, and inhibit metastasis, suggesting the potential of G6PD as a target for anti-tumor therapeutic strategies. Indeed, while challenges-including side effects-still remain, small-molecule G6PD inhibitors showing potential anti-tumor effect either when used alone or in combination with other anti-tumor drugs have been developed. This review provides an overview of the structural significance of G6PD, its role in and regulation of tumor development and progression, and the strategies explored in relation to G6PD-targeted therapy.

Design and synthesis of luotonin A-derived topoisomerase targeting scaffold with potent antitumor effect and low genotoxicity

Conventional topoisomerase (Topo) inhibitors typically usually exert their cytotoxicity by damaging the DNAs, which exhibit high toxicity and tend to result in secondary carcinogenesis risk. Molecules that have potent topoisomerase inhibitory activity but involve less DNA damage provide more desirable scaffolds for developing novel chemotherapeutic agents. In this work, we broke the rigid pentacyclic system of luotonin A and synthesized thirty-three compounds as potential Topo inhibitors based on the devised molecular motif. Further investigation disclose that two compounds with the highest antiproliferation activity against cancer cells, 5aA and 5dD, had a distinct Topo I inhibitory mechanism different from those of the classic Topo I inhibitors CPT or luteolin, and were able to obviate the obvious cellular DNA damage typically associated with clinically available Topo inhibitors. The animal model experiments demonstrated that even in mice treated with a high dosage of 50 mg/kg 5aA, there were no obvious signs of toxicity or loss of body weight. The tumor growth inhibition (TGI) rate was 54.3 % when 20 mg/kg 5aA was given to the T24 xenograft mouse model, and 5aA targeted the cancer tissue precisely without causing damage to the liver and other major organs.

Melittin kills A549 cells by targeting mitochondria and blocking mitophagy flux

ABSTRACT Melittin, a naturally occurring polypeptide found in bee venom, has been recognized for its potential anti-tumor effects, particularly in the context of lung cancer. Our previous study focused on its impact on human lung adenocarcinoma cells A549, revealing that melittin induces intracellular reactive oxygen species (ROS) burst and oxidative damage, resulting in cell death. Considering the significant role of mitochondria in maintaining intracellular redox levels and ROS, we further examined the involvement of mitochondrial damage in melittin-induced apoptosis in lung cancer cells. Our findings demonstrated that melittin caused changes in mitochondrial membrane potential (MMP), triggered mitochondrial ROS burst (Figure 1), and activated the mitochondria-related apoptosis pathway Bax/Bcl-2 by directly targeting mitochondria in A549 cells (Figure 2). Further, we infected A549 cells using a lentivirus that can express melittin-Myc and confirmed that melittin can directly target binding to mitochondria, causing the biological effects described above (Figure 2). Notably, melittin induced mitochondrial damage while inhibiting autophagy, resulting in abnormal degradation of damaged mitochondria (Figure 5). To summarize, our study unveils that melittin targets mitochondria, causing mitochondrial damage, and inhibits the autophagy-lysosomal degradation pathway. This process triggers mitoROS burst and ultimately activates the mitochondria-associated Bax/Bcl-2 apoptotic signaling pathways in A549 cells.

Abstract C108: Enhanced growth inhibition in lung cancer cells through NBF-006 combination therapy with chemotherapy or KRAS inhibitors

Abstract Background: Glutathione-S-transferase P (GSTP) is strongly up-regulated in many cancer types, specifically KRAS-mutant lung and is also a known regulator of oncogenic MAPK and PI3K pathways. In cancer cells, overexpression of GSTP regulates apoptosis by inhibiting Jun-p53 complex in JNK pathway. NBF-006 is a novel lyophilized lipid nanoparticle formulation, encapsulating a small interfering ribonucleic acid (siRNA) designed to inhibit the expression of GSTP in lung tumors. NBF-006 has been very well tolerated with early signs of efficacy in NSCLC patients in a phase 1 dose escalation and expansion study. As NBF-006 is well tolerated and unlikely to have drug-drug interactions, combination strategies are currently being developed. Due to the detoxifying role of GSTP, combination with chemo agents used to treat NSCLC may increase the sensitivity of cancer cells resulting in apoptosis and/or reduced resistance. As GSTP and KRAS enhance tumor growth and survival, inhibition of GSTP and KRAS may be synergistic. Methods: Anti-tumor activity of GSTP siRNA (NDT-05-1040), when combined with chemo agents, KRAS inhibitors or immune cells was assessed in a panel of KRAS mutant NSCLC lines. For chemo agents and KRAS inhibitors, NDT-05-1040 combinations was evaluated using Combenefit software and Loewe model. Results: Combination of NDT-05-1040 with chemotherapy drugs including paclitaxel, gemcitabine, cisplatin, and pemetrexed controlled tumor cell growth significantly better than either NBF-006 alone or the chemo treatment alone. In A549, H1373, H358, and H1792 lines, at low doses paclitaxel combined with NDT-05-1040 and resulted in synergistic cell killing (Loewe scores >20). Gemcitabine & Cisplatin combination with NDT-05-1040 in H1373 cells further improved cell killing by 5-fold (Loewe scores >20). Pemetrexed combined with NDT-05-1040 and led to additive cell killing in A549 (Loewe score >10). Combination of NDT-05-1040 with KRASG12C inhibitors, Sotorasib & Adagrasib were conducted in H358, H1222, and MiaPaca-2. Cell viability assays demonstrated that the combination resulted in strong synergistic, anti-proliferative effects at low doses in H358 cells (the greatest Loewe scores for Adagrasib and Sotorasib: 53 & 26, respectively). Conclusion: Results from the current studies suggest that NBF-006, a very well tolerated monotherapy, can be used in combination with other drugs currently indicated for KRAS-mutant NSCLC to achieve more potent antitumor effects. Citation Format: Cima Cina, Akinori Sugiyama, Murali Kuracha, Nazia Abbasi, Dominic Medina, Hiroyuki Tanaka, Sonya Zabludoff. Enhanced growth inhibition in lung cancer cells through NBF-006 combination therapy with chemotherapy or KRAS inhibitors [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr C108.

Abstract A052: Discovery and characterization of a p300-selective degrader demonstrates potent anti-tumor activity in preclinical models of prostate cancer

Abstract Strategies targeting androgen receptor (AR) in the treatment of prostate cancer (PrCa) have shown clear clinical benefit. However, a variety of AR-signaling bypass mechanisms ultimately result in progression to metastatic castrate resistant prostate cancer (mCRPC), and therapies for these patients remain an urgent unmet need. The histone acetyltransferase paralogs p300 and CBP, key coactivators of AR, are currently under investigation as attractive therapeutic targets to abrogate aberrant AR signaling but face a challenging clinical toxicity profile in hematopoietic cells. We postulate that p300 serves as a key regulator of AR signaling, and by sparing CBP we should retain potency against AR-driven PrCa while providing a larger safety margin. We discovered potent heterobifunctional degraders with demonstrated selectivity for p300 over CBP. Our tool compound exhibits degradation of p300 within 2 hours (DC50 = 7.5nM, Dmax = 79%) with minimal impact on CBP at 48 h (DC50 >1uM, Dmax = 5%). The AR/ARv7 driven VCaP PrCa cell line demonstrated exceptional sensitivity to this p300 degrader in proliferation assays (gIC50 = 0.8 nM) comparable to the dual p300/CBP inhibitor, CCS-1477, or the AR degrader, ARV-110 and undergoes a cytotoxic response. Profiled against known AR target genes, our degrader was able to suppress KLK2, KLK3 and FKBP5 transcript levels akin to CCS-1477 in both LNCaP and VCaP PrCa cell lines, supporting the hypothesis that p300, not CBP, acts as the primary coactivator of AR signaling. In addition, our compound led to greater downregulation of full-length AR, ARv7 and cMyc compared to CCS-1477. Subsequent assessment in an in vivo model (VCaP CDX) demonstrated tumor growth inhibition at the highest dose tested (88%; p=2.6e-4). Lastly, we profiled our p300 degrader in human bone marrow derived hematopoietic progenitor colony-forming assays where we observed a significant reduction in toxicity (IC50 = 3.9uM) compared to either CCS-1477 (IC50 = 121nM) or a dual degrader tool compound (IC50 = 16nM), supporting the hypothesis that a p300-specific mechanism will offer an improved therapeutic index. Treatment with our p300-specific degrader resulted in a potent anti-tumor effect on PrCa cells both in vitro and in vivo, comparing favorably to both dual p300/CBP inhibitor or AR degrader strategies. Furthermore, selective degradation of p300 has the potential to minimize toxicity to hematopoietic cells commonly observed with dual targeting of CBP and p300. Taken together, these results support the role p300 may play in driving AR-mediated signaling in PrCa and demonstrate that a p300-specific degrader may serve as an effective therapeutic modality in this disease setting. Citation Format: Mike Russell, Cassandra L Lowenstein, Xuqing Zhang, Jeremy Roach, Jianing Song, Rakesh Nagilla, Peter Orth, Matt Tudor, Qiaolin Deng, Zhihua Sui, Corey Strickland, Larry J Jolivette, E Scott Priestley, Helai P Mohammad. Discovery and characterization of a p300-selective degrader demonstrates potent anti-tumor activity in preclinical models of prostate cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A052.

Abstract C111: A novel human monoclonal antibody targeting active ADAM10 demonstrates anti-tumor effects in colorectal cancer

Abstract Colorectal cancer (CRC) is the third most diagnosed cancer in the USA and accounts for more than 600,000 deaths annually worldwide, primarily due to relapse with highly aggressive, chemo-resistant disease characterized by poorly differentiated cancer cells with stem cell-like properties. A common signature of these chemo-resistant tumors is dysregulation of Notch receptor signaling, as well as upregulation of its metalloprotease activator, ADAM10. Although small molecule inhibition of either Notch or ADAM10 has been shown to produce potent anti-tumor effects, these therapeutic strategies have failed in clinical trials primarily due to systemic toxicities, especially cytotoxic effects on the gastrointestinal tract and musculoskeletal system, highlighting the need for development of more targeted approaches. We have previously demonstrated that ADAM10 predominantly exists in an inhibited state in normal tissues but is activated in tumor cells through a conformational change in the extracellular domain, thus providing a potential target for tumor-specific modulation of ADAM10 activity. Here, we look to characterize a novel human monoclonal antibody agent (1H5) that selectively targets an exposed extracellular region of activated ADAM10 on tumor cells. We previously demonstrated that treatment with a murine version of the antibody (8C7), specific for the activated form of both mouse and human ADAM10, conferred a significant reduction in tumor burden against human CRC cell lines in cell culture and transplants in xenograft models, and relapse was prevented when 8C7 was combined with chemotherapy. Here, we demonstrate that the 1H5 monoclonal antibody effectively reduces cell viability in numerous colorectal cancer cell lines in vitro and induces marked tumor regression in established xenograft models through inhibition of Notch signaling. Through the use of additional genetic tools combined with in vitro and in vivo models, our preclinical study aims to fully evaluate 1H5 as novel therapeutic agent to treat aggressive colorectal cancer. We aim to propel this novel agent toward clinical-stage development. Citation Format: Pargol Mashati, Dan Sun, Eduardo Garcia Reino, Jessica A. Blandino, Ben Mason, Nayanendu Saha, Dimitar B. Nikolov, Prem Premsrirut. A novel human monoclonal antibody targeting active ADAM10 demonstrates anti-tumor effects in colorectal cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr C111.

Targeted Therapy of Central Nervous System Acute Lymphoblastic Leukemia with an Integrin α6-Targeted Self-Assembling Proapoptotic Nanopeptide

There is currently no effective targeted therapeutic strategy for the treatment of central nervous system acute lymphoblastic leukemia (CNS-ALL). Integrin α6 is considered a potential target for CNS-ALL diagnosis and therapy because of its role in promoting CNS-ALL disease progression. The targeted peptide D(RWYD) (abbreviated RD), with nanomolar affinity to integrin α6 was identified by peptide scanning techniques such as alanine scanning, truncation, and D-substitution. Herein, we developed a therapeutic nanoparticle based on the integrin α6-targeted peptide for treating CNS-ALL. The self-assembled proapoptotic nanopeptide D(RWYD)-D(KLAKLAK)2-GD(FFY) (abbreviated RD-KLA-Gffy) contains the integrin α6-targeted peptide RD, the well-known proapoptotic peptide D(KLAKLAK)2 (abbreviated KLA), and the self-assembling tetrapeptide GD(FFY) (abbreviated Gffy). The functional mechanism of RD-KLA-Gffy is clarified using different experiments. Our results demonstrate that RD-KLA-Gffy is highly enriched in CNS-ALL lesions and induces tumor cell apoptosis, thus reducing CNS-ALL disease burden and prolonging the survival of CNS-ALL mice without obvious toxicity. Moreover, the combined use of RD-KLA-Gffy and methotrexate (MTX) shows a potent antitumor effect in treating CNS-ALL, indicating that RD-KLA-Gffy plays an important role in suppressing CNS-ALL progression either as a single agent or in combination with MTX, which shows promise for application in CNS-ALL therapy.

Abstract C145: ANS014004, a novel small-molecule type II c-Met inhibitor effectively overcomes clinical-resistance MET mutations and exhibits antitumor activity in preclinical models of MET-amplified non-small cell lung cancer (NSCLC) and gastric cancer

Abstract Mesenchymal epithelial transition (MET) proto-oncogene receptor tyrosine kinase (RTK) is a cell surface receptor selective for hepatocyte growth factor (HGF), and is involved in embryogenesis regulation, wound healing, organ regeneration, angiogenesis, and immunomodulation. Aberrant MET oncogenic alterations include: MET exon 14 skipping (MET∆ex14) mutations; activating mutations in the kinase domain; MET gene amplification; MET fusions; and MET protein overexpression. These oncogenic alterations occur in a wide range of human solid cancers. Small-molecule type I c-Met inhibitors bind to the active (Asp-Phe-Gly “DFG-in” state) of c-Met within the adenosine triphosphate (ATP) binding pocket via the Y1230 residue, resulting in down-regulation of multiple pro-oncogenic downstream factors. Presently, type I c-Met inhibitors such as Capmatinib are used as monotherapies in patients with locally advanced or metastatic NSCLC with MET∆ex14 mutations. However, development of post-treatment resistance to type I c-Met inhibitors occurs clinically, including through acquired mutations in codons D1228 and Y1230. No drugs have been approved globally for MET alteration indications other than MET∆ex14 mutations. Next generation MET inhibitors are thus needed to treat patients harboring various MET oncogenic alterations beyond MET∆ex14, including post-treatment acquired mutations. ANS014004 is a newly developed small-molecule type II inhibitor of c-Met, binding to the inactive state of c-Met (“DFG-out”) in the ATP pocket. ANS014004 showed greater kinase inhibition activity against c-Met mutant kinases Y1230H and D1228A in vitro (IC50’s = 2.596 and 2.971 nM, respectively), when compared with Capmatinib, Cabozantinib, and Merestinib (IC50 values = >5000, ~10, and ~5 nM respectively). Further, ANS014004 showed a stronger anti-proliferative effect in MET mutant Ba/F3 cells TPR-Met D1228A and TPR-Met-Y1230H in vitro (IC50’s = 49.34 nM and 30.32 nM, respectively), when compared with Capmatinib (IC50 values>10000nM) and Merestinib (IC50 values: 37~100 nM). ANS014004 also showed potent antitumor effects in vitro in the human cancer tumor cell lines harboring various pathogenetic MET alterations: Hs-746T (gastric); MKN-45 (gastric); EBC-1 (NSCLC); NCI-H1993 (NSCLC); with IC50 ranging from 6.38~62.32 nM. In several subcutaneous xenograft tumor model studies using these same cell lines implanted in BALB/c nude mice, orally administered ANS014004 produced significant dose-dependent tumor growth inhibition, and the corresponding exposures of ANS014004 in plasma, tumor and brain samples increased with increasing dose. ANS0104001 was also observed to induce tumor regression in a MET-amplified intracranial xenograft model. Overall, the non-clinical studies of ANS014004 demonstrated it is a potent, orally-bioavailable type II c-Met inhibitor with activity against various pathogenetic MET alterations and with favorable absorption, distribution, pharmacokinetic, efficacy, and tolerability profiles in vivo. The molecule will enter a Phase 1 clinical study in the US soon. Citation Format: Gong Li, Hepeng Shi, Peilong Zhang, Xiangqiu Li, Wenli Lan, Linna Li, David Chung, Kevin Schaab. ANS014004, a novel small-molecule type II c-Met inhibitor effectively overcomes clinical-resistance MET mutations and exhibits antitumor activity in preclinical models of MET-amplified non-small cell lung cancer (NSCLC) and gastric cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr C145.

MRNA-HPV vaccine encoding E6 and E7 improves therapeutic potential for HPV-mediated cancers via subcutaneous immunization.

The E6 and E7 proteins of specific subtypes of human papillomavirus (HPV), including HPV 16 and 18, are highly associated with cervical cancer as they modulate cell cycle regulation. The aim of this study was to investigate the potential antitumor effects of a messenger RNA-HPV therapeutic vaccine (mHTV) containing nononcogenic E6 and E7 proteins. To achieve this, C57BL/6j mice were injected with the vaccine via both intramuscular and subcutaneous routes, and the resulting effects were evaluated. mHTV immunization markedly induced robust T cell-mediated immune responses and significantly suppressed tumor growth in both subcutaneous and orthotopic tumor-implanted mouse model, with a significant infiltration of immune cells into tumor tissues. Tumor retransplantation at day 62 postprimary vaccination completely halted progression in all mHTV-treated mice. Furthermore, tumor expansion was significantly reduced upon TC-1 transplantation 160 days after the last immunization. Immunization of rhesus monkeys with mHTV elicited promising immune responses. The immunogenicity of mHTV in nonhuman primates provides strong evidence for clinical application against HPV-related cancers in humans. All data suggest that mHTV can be used as both a therapeutic and prophylactic vaccine.

A Phase 1 Trial of OPB-111077 in Combination with Bendamustine (B) and Rituximab (R) in Patients with Relapsed or Refractory (R/R) Diffuse Large B-Cell Lymphoma (DLBCL): Preliminary Results of Dose-Escalation Stage

Background OPB-111077, a first-in-class small molecule, is an orally available antitumor agent with an inhibitory effect on mitochondrial oxidative phosphorylation (OXPHOS). OPB-111077 inhibits mitochondrial respiratory chain complex I, which leads to inhibition of energy production and activation of the AMPK-mTOR energy stress sensor pathway, as well as inhibition of the signal transducer and activator of transcription (STAT) 3 pathway. In non-clinical studies using cultured cells and murine xenograft models, OPB-111077 showed antitumor effects in a variety of tumor cell lines, including hematological and solid tumors, and OPB-111077 in combination with an alkylating agent has a potential synergistic antitumor effect for malignant lymphoma. Here we report the preliminary results from the dose-escalation (DE) stage of an ongoing phase 1 trial (NCT04049825) of OPB-111077 in combination with bendamustine (B) and rituximab (R) in patients with relapsed or refractory (R/R) diffuse large B-cell lymphoma (DLBCL). Method This trial is composed of 2 stages. OPB-111077 is administered in combination with B in a DE stage and with B and R in a dose-expansion (EX) stage. The DE stage uses a 3+3 design. The objectives of the DE stage are to evaluate the tolerability and safety of OPB-111077 in combination with B and to determine the recommended dose for the EX stage based on dose-limiting toxicity (DLT). A secondary objective is to evaluate efficacy. Response assessment is based on IWG Lugano response criteria for non-Hodgkin lymphoma (NHL) (2014). Key eligibility criteria include age 20 to 80 years (inclusive), at least initial standard treatment, measurable lesions, and Eastern Cooperative Oncology Group performance status score of 0 or 1. OPB-111077 is orally administered once daily on Days 3 to 6, Days 10 to 13, and Days 17 to 20 of each cycle. The duration of each cycle is 21 days, except for Cycle 1, which is 23 days. The dose in the first cohort is 200 mg/day, increasing as appropriate to 400 mg/day in Cohort 2 and then to 600 mg/day in Cohort 3. B is administered intravenously at 120 (or 90) mg/m 2 once daily on Days 1 and 2 of each cycle for up to 6 cycles. Results As of 31 Jan 2023, 16 patients had been treated with OPB-111077 + B (120 mg/m 2) at OPB-111077 doses of 200 mg (Cohort 1, n = 3), 400 mg (Cohort 2, n = 6), and 600 mg (Cohort 3, n = 7). Patients had received 1 to 6 prior lines of chemotherapy, median age was 70.0 (range 54 - 79) years, and 56.3% were male. Median treatment duration was 69.0 days. No DLT was observed in Cohort 1 or in the first 3 patients in Cohort 2. In Cohort 3, 2 patients experienced DLTs of neutropenia (grade 4) and neutropenia (grade 3, dose interruption of ≥21 consecutive days), respectively. As the Efficacy and Safety Evaluation Committee (ESEC) judged the OPB-111077 600-mg dose to be intolerable, in accordance with the scheme for the DE stage, new 3 patients were added to Cohort 2. One of the 6 patients in Cohort 2 experienced DLTs of anorexia (grade 3) and neutropenia (grade 4, dose interruption of ≥21 consecutive days). Based on the ESEC's recommendation, the recommended dose for the EX stage was set at OPB-111077 400 mg + B 120 mg/m 2. The most common (≥90%) treatment-related adverse events of any grade were lymphopenia (93.8%, grade 3: ≥93.8%), leukopenia (93.8%, grade 3: ≥81.3%), and nausea (93.8%, grade 3: ≥18.8%). The overall response rate [complete response (CR) + partial response (PR)] and CR rate were respectively 62.5% and 43.8%. Median progression-free survival was 257.0 days. Conclusion OPB-111077 400 mg + B 120 mg/m 2 combination treatment showed a tolerable safety profile. The preliminary data are encouraging, especially regarding the efficacy of OPB-111077 combination treatment, including CR in patients with DLBCL. The DE stage of the trial is still ongoing and the EX stage, which will evaluate the safety and efficacy of OPB-111077 in combination with B + R, is now recruiting.

Pyroptosis-related gene signature: A predictor for overall survival, immunotherapy response, and chemosensitivity in patients with pancreatic adenocarcinoma

BackgroundPancreatic adenocarcinoma (PAAD) is a lethal malignancy with high levels of heterogeneity. Pyroptosis is thought to influence the development of various tumors. Nevertheless, the role of pyroptosis-related genes (PRGs) in prognostic risk stratification and therapeutic guidance for PAAD remains ambiguously. MethodsTranscriptome profile and clinical information of PAAD patients were retrieved from The Cancer Genome Atlas (TCGA) as well as Gene Expression Omnibus (GEO) databases, followed by differential analysis. Patients were divided into distinct pyroptosis phenotype subtypes based on the characteristic of differently expressed PRGs (DEPRGs). Then a PRG signature was established through univariate analysis and LASSO algorithm in the training set to assess the prognostic risk, and its reliability was verified in the validation set using receiver operating characteristic(ROC) curve. The correlation of risk score with tumor microenvironment(TME), TMB and chemotherapeutic drug sensitivity were also analyzed. In addition, a nomogram was constructed to promote better clinical application. ResultsA total of 28 DEPRGs were determined in the integrated TCGA-GEO datasets. Patients were divided into three pyroptosis phenotype subtypes, Kaplan-Meier curve suggested patients in cluster B had a worse prognosis than those in cluster A and C. Then a price signature comprised of 8 PRGs was generated. TME analysis suggested that the low-risk subgroup displayed potential stronger antitumor immune effect and might respond better to immune checkpoint inhibitors (ICIs) therapy. Furthermore, PRG signature exhibited favorable discriminatory ability for TMB status and the sensitivity of multiple conventional chemotherapeutic agents including paclitaxel. Ultimately, we constructed a promising nomogram according to the risk score and N stage with good predictive accuracy compared with the actual overall survival (OS) probabilities. ConclusionWe established an 8-gene signature that could be regarded as an independent prognostic risk factor for PAAD patients. The 8-gene signature could provide rationale for immunotherapy and chemotherapy, which might help clinicians make precise individualized treatment regimens.

Efficacy of Protac-Based EZH2 Degrader MS1943 in Lymphoid Malignancies: A Comprehensive Study on the Combined Effects of MS1943 and Ibrutinib

Introduction: This study delves into the therapeutic potential of the Enhancer of zeste homolog 2 (EZH2) degrader MS1943 and the Bruton's tyrosine kinase (BTK) inhibitor Ibrutinib in B-cell lymphomas. The study's primary objective was to compare the efficacy of MS1943, a Proteolysis-targeting chimera (PROTAC)-based degrader, with the FDA-approved EZH2 inhibitor Tazemetostat. The study also aimed to explore the potential synergistic effects of combining these therapies with Ibrutinib, a widely used BTK inhibitor in lymphoma treatment. Materials and Methods: The study employed a range of lymphoma cell lines, including B-cell and T-cell lymphomas, which were treated with MS1943, Tazemetostat, and Ibrutinib, either alone or in combination. The impact of these treatments on cell viability was evaluated using the Cell Counting Kit-8 (CCK-8) assay at different time points and concentrations. Cell cycle distribution was assessed using flow cytometric analysis, and the expression of downstream effectors of the unfolded protein response (UPR) pathway, including Xbp1, Chop, and Bip, was examined using quantitative polymerase chain reaction (qPCR). Western blot analysis was performed to investigate the association with specific apoptosis-related pathways. Results: The results demonstrated that MS1943 exhibited significantly greater inhibitory effects than Tazemetostat in all lymphoma cell lines tested, particularly in B-cell lymphomas. MS1943 treatment resulted in increased expression of UPR pathway effectors, suggesting a correlation between the observed anti-proliferative effects and the differential expression of these factors. The combination of MS1943 and Ibrutinib demonstrated significant inhibitory effects across all B-cell lymphoma cell lines, with the most prominent cytotoxicity observed after 72 hours of culture. The combination treatment resulted in an increase in the number of cells in the G2/M phase, accompanied by a decrease in the number of cells in the G0/G1 phase. Furthermore, the combination treatment led to a notably higher proportion of apoptotic and necroptotic cells compared to either individual treatment or the combination of Ibrutinib and Tazemetostat. The combination treatment resulted in an increase in the expression of p53 and PUMA, and a significant increase in miR29b expression. Discussion: Our findings suggest that the simultaneous treatment of MS1943 and Ibrutinib in B-cell lymphoma induces apoptosis through the miR29b-mediated p53 upregulated pathway, thereby exhibiting potent anti-tumor effects. The study provides compelling evidence that MS1943 exhibits a pronounced inhibitory effect on B-cell lymphomas compared to Tazemetostat, highlighting its potential as a promising therapeutic agent in the treatment of B-cell lymphomas. The combination of MS1943 and Ibrutinib enhances apoptotic and necroptotic cell death, resulting in a significant reduction in viable cells. This study provides valuable insights into the mechanisms underlying the anti-tumor effects of this drug combination, highlighting the potential of MS1943 as an effective treatment option for lymphoma.

Preclinical Evaluation of the Efficacy of Human Sirpα Antibodies for B-Cell Lymphoma Immunotherapy in Humanized Mouse Models

Tumor-associated macrophages (TAMs) are abundant in the tumor microenvironment and are considered potential targets to enhance cancer immunotherapy against B-cell lymphomas. However, faithful and predictive preclinical tumor models of human B-cell lymphomas for the evaluation of therapeutics targeting human TAMs have not been established yet. To examine the antitumor effects of agents targeting human TAMs in vivo, we here established preclinical tumor xenograft models based on Rag2-/-Il2rg-/- immunodeficient mice that express multiple human cytokines, such as M-CSF, IL-3, GM-CSF, and THPO (MITRG mice), and have been reconstituted with a human immune system (HIS) by transplantation of human CD34+ hematopoietic stem and progenitor cells (HIS-MITRG mice). HIS-MITRG mice supported the growth of both the human cell line (Raji)-derived and diffuse large B cell lymphoma (DLBCL) patient-derived xenograft tumors as well as the infiltration of human macrophages into these tumors. We examined the potential antitumor effect of an antibody to human SIRPα (SE12C3) that inhibits the interaction of CD47 on tumor cells with SIRPα on human macrophages and thereby promotes Fcγ receptor-mediated phagocytosis of tumor cells by human macrophages. Combined treatment with rituximab and SE12C3 inhibited Raji tumor growth in HIS-MITRG mice to a markedly greater extent than did rituximab monotherapy in a therapeutic model started after the engraftment of the tumor. This enhanced antitumor effect was dependent on human macrophages and attributable to enhanced rituximab-dependent phagocytosis of lymphoma cells by human macrophages. Treatment with rituximab and SE12C3 also induced reprogramming of human TAMs towards a proinflammatory phenotype. Furthermore, the combination treatment essentially prevented the growth of DLBCL patient-derived xenograft tumors in HIS-MITRG mice. Our findings thus support the utility of HIS-MITRG mice as a model for the preclinical in vivo evaluation of potential therapeutics, such as antibodies to human SIRPα, that target human TAMs.

Recombinant Production of TP4-LYC1, A New Chimeric Peptide with Targeted Cytotoxicity to HeLa Cells.

Tilapia Piscidin 4 (TP4) showed potential anti-tumor effects against various cancer cells. Lycosine-1 (LYC1), is another Antimicrobial Peptides (AMP) from spider venom with targeted penetration to cancer cells without any adverse effects on normal cells. The aim of this study was to produce a soluble recombinant fusion peptide in order to diminish the cytotoxicity of TP4 against normal cells. In order to express of TP4-LYC-1, TP4, and LYC1 in fusion to the inteins1/2 of pTWIN-1 vector, induction condition was optimized to earn soluble peptides. Auto-cleavage induction of inteins1/2 was performed based on IMPACT® manual and their effect on cell viability of HeLa and HUVEC cells was surveyed by MTT assay. The best condition for accessing the most soluble peptide in fusion to the inteins was approximately similar for all three peptides (0.1 mM of IPTG, at 22°C). After the induction of self-cleavage of inteins, a band in 3, 3, and 6 kDa was observed on tricine-SDS-PAGE. The IC50 values of TP4-LYC1 and TP4 against HeLa cells were calculated as 0.83, and 2.75 μM, respectively. In the present study, a novel chimeric peptide, TP4-LYC1, was successfully produced. This fusion protein can act as a safe bio-molecule with potent cytotoxic effects against cancer cells, but the penetration ability and determination of cell death mechanism must be performed in order to have more precise view on the apoptosis induction of this recombinant peptide.