Pal, Krishnendu

Optineurin (OPTN) is a multifunctional adaptor protein that regulates diverse cellular processes, including inflammatory signaling, autophagy, vesicular trafficking, and immune responses. This multifaceted role of OPTN is made possible by the presence of a complex structure comprising multiple domains that interact with different proteins to exert various functions important for modulating key signaling processes. Mutations in OPTN are linked with several human pathologies including glaucoma, Paget’s disease of bone, Crohn’s disease, and neurodegenerative diseases such as amyotrophic lateral sclerosis, and dementia. Emerging evidence suggests that OPTN has a complex and context-dependent role in cancer biology as well. It is upregulated in pancreatic ductal adenocarcinoma and hepatocellular carcinoma but downregulated in lung and colorectal cancers, indicating its dual role as a potential oncogene or tumor suppressor depending on the cellular environment. Additionally, OPTN plays a critical role in preventing immune evasion in colorectal cancer by maintaining interferon-gamma receptor 1 (IFNGR1) expression and supporting dendritic cell-mediated T-cell priming, thereby enhancing antitumor immune responses. Despite its significance in oncogenic pathways and immune regulation, the therapeutic potential of targeting OPTN in cancer remains largely unexplored. This review aims to provide a comprehensive understanding of OPTN’s pleiotropic functions, highlighting its role in autophagy, inflammation, immune surveillance, and cancer progression. By elucidating its diverse regulatory mechanisms, we seek to encourage further research into the therapeutic implications of OPTN in cancer treatment and immunotherapy.

Abstract Background Glioblastoma (GBM) is the most common high-grade primary malignant brain tumor, characterized by a notably poor prognosis. Current treatments for GBM have shown limited effectiveness in improving patient survival, highlighting the urgent need for novel therapeutic strategies. Combination therapy offers significant potential in overcoming resistance by targeting multiple signaling pathways; however, it often comes with increased toxicity compared to monotherapy. Co-encapsulating multiple therapeutic agents into a tumor-targeted drug delivery platform holds promise for overcoming these limitations and improving treatment outcomes. Methods We developed a tumor-targeted liposomal nanoformulation (TTL) using phospholipids, cholesterol, DSPE-(PEG)2000-OMe, and a proprietary tumor-targeting peptide (TTP). The TTL was loaded with everolimus (TTL-E), vinorelbine (TTL-V), rapamycin (TTL-R), a combination (TTL-EV), or (TTL-RV). These formulations were tested in vivo on orthotopic GBM mice, combined with temozolomide and radiation. RNA sequencing was performed to identify molecular and transcriptome changes post-treatment. Results TTL demonstrated tumor-specific uptake, effectively delivering drugs to GBM tumors. TTL-EV and TTL-RV outperformed single-drug formulations. Radiation combined with TTL-EV/RV improved tumor growth inhibition and survival, while temozolomide provided minimal benefit. Transcriptome analysis revealed differentially expressed genes (DEGs) linked to DNA damage repair, cell cycle, metabolism, and extracellular matrix pathways. Conclusion TTL crossed the blood-brain barrier, targeting tumors effectively. Radiation plus TTL-EV/RV enhanced tumor suppression and survival in GBM models. Gene expression analysis identified DEGs related to DNA damage and cell death. Mechanistic studies suggest TTL-EV plus radiation inhibits mTOR/MAPK pathways and sensitizes tumors to radiation. These findings offer a potential approach for improving GBM treatment.

Background/Objectives: Pancreatic ductal adenocarcinoma (PDAC) remains one of the most lethal malignancies, characterized by aggressive tumor biology, poor vascularization, dense stromal barriers, and profound resistance to chemotherapy. GAIP-interacting protein C-terminus 1 (GIPC1), a PDZ-domain-containing adaptor protein, is highly overexpressed in PDAC and plays a critical role in tumor progression and chemoresistance. This study aimed to develop and evaluate a novel tumor-targeted liposomal siRNA delivery system (LGIPCsi) to silence GIPC1 and enhance the therapeutic efficacy of gemcitabine (GEM) in PDAC; Methods: LGIPCsi nanoparticles were synthesized and optimized for physicochemical stability, siRNA complexation efficiency, and tumor-targeting capability. Their therapeutic efficacy was assessed using in vitro pancreatic cancer cell models and in vivo orthotopic and patient-derived xenograft (PDX) models of PDAC. Biodistribution, tumor uptake, and antitumor efficacy were evaluated following systemic administration. Combination studies were performed to assess the synergistic effects of LGIPCsi and GEM; Results: GIPC1 silencing significantly sensitized pancreatic cancer cells to GEM, resulting in enhanced inhibition of tumor cell proliferation in vitro. In vivo, systemic administration of LGIPCsi achieved efficient intratumoral delivery of siGIPC1, leading to marked tumor growth suppression. Combination therapy with GEM and LGIPCsi produced synergistic antitumor effects, with substantial tumor regression compared to monotherapy groups. Importantly, no significant systemic toxicity was observed in treated animals; Conclusions: This study identifies GIPC1 as a promising therapeutic target in PDAC and demonstrates that tumor-targeted siRNA nanomedicine can effectively overcome chemoresistance when combined with standard chemotherapy. The LGIPCsi platform offers a rational and translational strategy to enhance treatment efficacy in PDAC through targeted RNAi-based combination therapy.

Abstract Background Renal cell carcinoma (RCC) was historically considered to be less responsive to radiation therapy (RT) compared to other cancer indications. However, advancements in precision high-dose radiation delivery through single-fraction and multi-fraction stereotactic ablative radiotherapy (SABR) have led to better outcomes and reduced treatment-related toxicities, sparking renewed interest in using RT to treat RCC. Moreover, numerous studies have revealed that certain therapeutic agents including chemotherapies can increase the sensitivity of tumors to RT, leading to a growing interest in combining these treatments. Here, we developed a rational combination of two radiosensitizers in a tumor-targeted liposomal formulation for augmenting RT in RCC. The objective of this study is to assess the efficacy of a tumor-targeted liposomal formulation combining the mTOR inhibitor everolimus (E) with the survivin inhibitor YM155 (Y) in enhancing the sensitivity of RCC tumors to radiation. Experimental Design: We slightly modified our previously published tumor-targeted liposomal formulation to develop a rational combination of E and Y in a single liposomal formulation (EY-L) and assessed its efficacy in RCC cell lines in vitro and in RCC tumors in vivo. We further investigated how well EY-L sensitizes RCC cell lines and tumors toward radiation and explored the underlying mechanism of radiosensitization. Results EY-L outperformed the corresponding single drug-loaded formulations E-L and Y-L in terms of containing primary tumor growth and improving survival in an immunocompetent syngeneic mouse model of RCC. EY-L also exhibited significantly higher sensitization of RCC cells towards radiation in vitro than E-L and Y-L. Additionally, EY-L sensitized RCC tumors towards radiation therapy in xenograft and murine RCC models. EY-L mediated induction of mitotic catastrophe via downregulation of multiple cell cycle checkpoints and DNA damage repair pathways could be responsible for the augmentation of radiation therapy. Conclusion Taken together, our study demonstrated the efficacy of a strategic combination therapy in sensitizing RCC to radiation therapy via inhibition of DNA damage repair and a substantial increase in mitotic catastrophe. This combination therapy may find its use in the augmentation of radiation therapy during the treatment of RCC patients.

Abstract Purpose: Although renal cell carcinoma (RCC) is relatively more resistant to radiation therapy (RT) compared to other cancer indications, recent advances in single-fraction and multi-fraction stereotactic ablative radiotherapy (SABR) made it possible to achieve better outcome and reduce treatment related toxicities. In addition, with studies demonstrating that certain therapeutic agents including chemotherapies can act as radiosensitizers, the idea of combining radiotherapy with those agents are being actively pursued. Among them, mammalian target of rapamycin (mTOR) inhibitor everolimus (E) and survivin inhibitor YM155 (Y) are of special interest since they target multiple RT-resistance mechanisms simultaneously. Additionally, YM155 has been shown to help in overcoming resistance to mTOR inhibitors in various cancer including RCC. Hence, the present study aims to evaluate the efficacy of a tumor-targeted liposomal formulation combining everolimus and YM155 (EY-L) in sensitizing RCC tumors towards radiation. Experimental Design: Previously we developed a tumor-targeted liposomal formulation and used it successfully to deliver multiple drugs for the treatment of RCC in both immune-deficient and immune-competent mouse models. Here we developed a similar formulation combining everolimus and YM155 (EY-L) and assessed its efficacy in RCC cell lines in vitro and in RCC tumors in vivo. We further evaluated the efficacy of EY-L in sensitizing RCC cell lines and tumors towards radiation in vitro using clonogenic assay and in vivo in RCC xenografts in immunodeficient mice and murine RCC models in immunocompetent mice. Results: EY-L (2 mg/kg E, 1.4 mg/kg Y, twice a week intravenous administration) was better than E-L or Y-L, the corresponding single drug loaded formulations in containing the primary tumor growth and improving survival in an immunocompetent syngeneic mouse model of RCC. Furthermore, EY-L leads to significantly higher sensitization of RCC cells towards radiation in vitro than E-L or Y-L. Finally, EY-L was able to sensitize RCC tumors towards radiation therapy in both RCC xenograft and murine RCC model. Conclusion: Taken together, our study demonstrated the efficacy of a strategic combination therapy in sensitizing RCC to radiation therapy using in vitro clonogenic assay and in vivo radiosensitization studies in multiple RCC tumor models. Citation Format: Krishnendu Pal, Vijay S. Madamsetty, Hari K. Rachamala, Shamit K. Dutta, Enfeng Wang, Debabrata Mukhopadhyay. Combined inhibition of mammalian target of rapamycin (mTOR) and survivin augments radiation therapy in renal cell carcinoma [abstract]. In: Proceedings of the AACR Special Conference: Advances in Kidney Cancer Research; 2023 Jun 24-27; Austin, Texas. Philadelphia (PA): AACR; Cancer Res 2023;83(16 Suppl):Abstract nr B026.

Despite recent advancements, effective treatment for pancreatic ductal adenocarcinoma (PDAC) has remained elusive. The overall survival rate in PDAC patients has been dismally low due to resistance to standard therapies. In fact, the failure of monotherapies to provide long-term survival benefits in patients led to ascension of several combination therapies for PDAC treatment. However, these combination therapies provided modest survival improvements while increasing treatment-related adverse side effects. Hence, recent developments in drug delivery methods hold the potential for enhancing therapeutic benefits by offering cocktail drug loading and minimizing chemotherapy-associated side effects. Nanoformulations-aided deliveries of anticancer agents have been a success in recent years. Yet, improving the tumor-targeted delivery of drugs to PDAC remains a major hurdle. In the present paper, we developed several new tumor-targeted dual intervention-oriented drug-encapsulated (DIODE) liposomes. We successfully formulated liposomes loaded with gemcitabine (G), paclitaxel (P), erlotinib (E), XL-184 (c-Met inhibitor, X), and their combinations (GP, GE, and GX) and evaluated their in vitro and in vivo efficacies. Our novel DIODE liposomal formulations improved median survival in comparison with gemcitabine-loaded liposomes or vehicle. Our findings are suggestive of the importance of the targeted delivery for combination therapies in improving pancreatic cancer treatment.

Schematic representation demonstrating the fabrication and in vivo evaluation of an immune-modulatory nano-formulation consisting of irinotecan and curcumin in immune-competent mouse models of pancreatic adenocarcinoma.

Abstract Quiescin sulfhydryl oxidase 1 (QSOX1) is an enzyme overexpressed by many different tumor types. QSOX1 catalyzes the formation of disulfide bonds in proteins. Because short hairpin knockdowns (KD) of QSOX1 have been shown to suppress tumor growth and invasion in vitro and in vivo, we hypothesized that chemical compounds inhibiting QSOX1 enzymatic activity would also suppress tumor growth, invasion, and metastasis. High throughput screening using a QSOX1-based enzymatic assay revealed multiple potential QSOX1 inhibitors. One of the inhibitors, known as “SBI-183,” suppresses tumor cell growth in a Matrigel-based spheroid assay and inhibits invasion in a modified Boyden chamber, but does not affect viability of nonmalignant cells. Oral administration of SBI-183 inhibits tumor growth in 2 independent human xenograft mouse models of renal cell carcinoma. We conclude that SBI-183 warrants further exploration as a useful tool for understanding QSOX1 biology and as a potential novel anticancer agent in tumors that overexpress QSOX1.

Abstract Resistance to chemotherapy remains a major obstacle to improving outcomes of cancer treatment. Recent data suggest intrinsic functions of immune checkpoint proteins, including PD-L1, that may be distinct from their immunoregulatory roles. We found that colorectal cancer (CRC) cells with mutant BRAF(V600E) or KRASalleles show increased PD-L1 transcripts and expression of cell surface PD-L1 protein compared to those with wild-type copies. Furthermore, ectopic expression of BRAFV600E or mutant KRAS induced PD-L1 whose upregulation was attenuated by inhibition of RAS-RAF-MEK-ERK signaling. Combined knockdown of MEK/ERK effectors c-JUN and YAP markedly reduced PD-L1. Knockout of PD-L1 in RKO cells resulted in a reduction in chemotherapy-induced DNA double strand breaks (pH2AX) and caspase-3 cleavage compared to parental cells. Results were confirmed in PD-L1 knockout MC38 murine CRC cells where re-expression of PD-L1, but not its deletion mutants, was shown to induced pH2AX and apoptosis. In cells with knockout of PD-L1, reductions in p-AKT and the BH3-only proteins BIM and BIK were observed that could be restored by re-expression of PD-L1. Treatment of cells with an anti-PD-L1 antibody reduced p-AKT, BIM and BIK, and also attenuated chemotherapy-induced apoptosis. Re-expression of BIM in PD-L1 knockout cells restored apoptosis. Murine tumor xenografts generated from cells with knockout of PD-L1 displayed resistance to oxaliplatin-induced tumor regression compared to tumors derived from parental cells. Using TCGA datasets, human CRCs with high vs low PD-L1 mRNA levels were significantly associated with better survival. In summary, we identified a non-immune function of tumor cell-intrinsic PD-L1 that upregulates BH3-only proteins BIM and BIK to promote apoptosis, thereby suggesting its potential as a predictive biomarker. Citation Format: Daofu Feng, Krishnendu Pal, Debabrata Mukhopadhyay, Xin Liu, Haidong Dong, Shengbing Huang, Frank A. Sinicrope. Tumor intrinsic PD-L1 is mediated by BRAFV600E and can regulate drug-induced apoptosis in human colon cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 705.