Molecular Cancer Therapeutics最新文献

Colorectal cancer (CRC) is a leading cause of cancer-related deaths worldwide. The current standard of care for patients may involve surgery, chemotherapy, and immune checkpoint inhibitors (ICIs), but these approaches typically fail to secure durable responses against late-stage disease. Regorafenib (REG) is an FDA-approved tyrosine kinase inhibitor with immunomodulating properties for CRC patients who progress on standard care, but 5-year relative survival rates for individuals dosed with the drug as a monotherapy are poor. We hypothesize that REG may be more appropriately leveraged alongside immunotherapeutic agents that specifically stimulate T cell infiltration and activation within the tumor microenvironment (TME). We engineered a PD-L1/CD3 bispecific antibody (bsAb) that simultaneously binds PD-L1-expressing CRC cells and stimulates activated T cells in order to investigate combination strategies with REG in pre-clinical models of CRC. Combined REG + bsAb therapy safely initiated and sustained inhibition against MC38 and CT26 progression in vivo, and these effects correlated to improved CD8+ T cell infiltration and activity within a Type-1-prone TME. Additionally, cytotoxic CD8+ T cells from REG + bsAb-sensitized mice exhibited heightened tumor cell reactivity compared to animals treated with either agent alone. Therefore, the immunomodulatory benefits of REG can be effectively paired with a bsAb that anchors to CRC cells, diminishes immunosuppression (through PD-L1 blockade), and activates/sustains antigen-specific CD8+ T cells within the TME. Our newly described REG + bsAb regimen led to improved anti-tumor outcomes pre-clinically and may represent a promising future approach for CRC patients.

The limitations of first-generation antibody-drug conjugate (ADC) technologies include suboptimal stability and efficacy, poor safety profiles, and challenging manufacturing processes. In this study, we describe an anti-CD79b-monomethyl auristatin E (MMAE) ADC generated using a novel peptide-based linker technology that allows for site-specific linker-payload conjugation to native antibodies in only one step. The ADC comprises native polatuzumab as the targeting antibody and a linker-payload consisting of a RKAA-peptide linker and MMAE. We compared our anti-CD79b-RKAA-MMAE ADC with polatuzumab vedotin (PV), the FDA-approved ADC for diffuse large B-cell lymphoma. In the clinic, PV shows significant instability in circulation, leading to strong and dose-limiting side effects, including neutropenia and peripheral neuropathy. The anti-CD79b-RKAA-MMAE ADC showed optimal biophysical properties with a well-defined drug-to-antibody ratio of 2. It demonstrated potent cytotoxicity in multiple cancer cell lines and was very stable in mouse, cynomolgus monkey, and human sera. The anti-CD79b-RKAA-MMAE conjugate showed equal antitumor efficacy at half the payload dose compared with PV in different xenograft models. At equal MMAE concentrations, greater tumor growth inhibition and a considerably longer duration of response were observed. Ultimately, the highest nonseverely toxic dose of 30 mg/kg was determined in a 4-week repeat-dose toxicology study in rats, which is a 3-fold higher ADC dose than reported for PV. In summary, the data show that our novel site-specific bioconjugation technology enabled the generation of an anti-CD79b-RKAA-MMAE ADC with highly favorable biophysical properties and a greatly improved therapeutic index by a factor of 4 to 6 compared with PV. The ADC may therefore represent a safe and efficacious alternative for patients with diffuse large B-cell lymphoma.

Fibroblast activation protein (FAP) is overexpressed on cancer-related fibroblasts (CAFs), making it an important target for cancer diagnosis and treatment, but limited tumor retention hinders late-stage diagnosis and radionuclide therapy. In this study, three albumin-bound FAPI radioligands, 68Ga/177Lu-DOTA-ALB-01, 68Ga/177Lu-DOTA-ALB-02, and 68Ga/177Lu-DOTA-ALB-03, were synthesized and evaluated for their in vitro stability, binding affinity, in vivo biodistribution, and tumor uptake using 68Ga and 177Lu labeling. All radioligands are stable in saline and plasma and exhibit high FAP binding affinity. 177Lu-DOTA-ALB-02 has longer retention in circulation than 177Lu-FAPI-46 and other radioligands. Continuous tumor accumulation was observed during imaging with both 177Lu-DOTA-ALB-01 and 177Lu-DOTA-ALB-02. Notably, 177Lu-DOTA-ALB-02 had a significant tumor/ nontarget (T/NT) ratio as indicated by biodistribution data. The outstanding tumor retention properties of 177Lu-DOTA-ALB-02 have been demonstrated in small animal single photon emission computed tomography (micro-SPECT) imaging and biodistribution studies, therefore it is considered the albumin-binding FAPI with the most favorable pharmacokinetic and imaging properties, worthy of further clinical investigation.

Precision medicine and genomic profiling with target-specific therapy directed to cancer cell receptors have improved the outcome of many recalcitrant cancers. Strategies to deliver gene therapy to downregulate cancer driver genes have been challenging in vivo. Pancreatic cancer has the poorest survival of all solid tumors due to the lack of target-specific therapies and its characteristic tumor microenvironment with dense fibrosis and abundant immunosuppressive M2-polarized macrophages. In this study, we designed a panel of locked nucleic acid (LNA) gapmer antisense oligonucleotides (ASO) directed to human gastrin mRNA. We tested their efficacy by downregulation of mRNA and growth inhibition in vitro. The most effective, Gapmer-90, was modified for in vivo therapeutics by thiol-maleimide click chemistry to render it target-specific to the CCK-B receptor. This G-protein coupled receptor is over-expressed in pancreatic cancers. Mice bearing orthotopic human pancreatic tumors were treated with PBS (control), an untargeted gapmer, or receptor-targeted gapmers at low (60nM) and high (120nM) concentrations. Uptake of the gapmer was measured in tissues using a complementary probe. We found that the receptor-targeted gapmer significantly enhanced uptake in vivo and decreased growth and metastases of human pancreatic tumors in a dose-related fashion without off-target toxicity. The targeted-specific gapmer also altered the tumor microenvironment by decreasing fibrosis and reducing M2-polarized macrophages. Collectively, our results provide evidence that LNA gapmers are a unique tool to deliver antisense oligonucleotides for therapy to recalcitrant cancers. Rendering the gapmers target-specific allows for selective uptake by receptor internalization, improving efficacy, and decreasing off-target toxicity.

KRASG12C selective inhibitors, such as sotorasib and adagrasib, have raised hopes of targeting other KRAS-mutant alleles in patients with cancer. We report that KRAS wild-type (WT)-amplified tumor models are sensitive to treatment with the small-molecule KRAS inhibitors BI-2493 and BI-2865. These pan-KRAS inhibitors directly target the "OFF" state of KRAS and result in potent antitumor activity in preclinical models of cancers driven by KRAS-mutant proteins. In this study, we used the high-throughput cellular viability Profiling Relative Inhibition Simultaneously in Mixtures assay to assess the antiproliferative activity of BI-2493 in a 900+ cancer cell line panel, expanding on our previous work. KRAS WT-amplified cancer cell lines, with a copy number >7, were identified as the most sensitive, across cell lines with any KRAS alterations, to our pan-KRAS inhibitors. Importantly, our data suggest that a KRAS "OFF" inhibitor is better suited to treat KRAS WT-amplified tumors than a KRAS "ON" inhibitor. KRAS WT amplification is common in patients with gastroesophageal cancers in which it has been shown to act as a unique cancer driver with little overlap to other actionable mutations. The pan-KRAS inhibitors BI-2493 and BI-2865 show potent antitumor activity in vitro and in vivo in KRAS WT-amplified cell lines from this and other tumor types. In conclusion, this is the first study to demonstrate that direct pharmacologic inhibition of KRAS shows antitumor activity in preclinical models of cancer with KRAS WT amplification, suggesting a novel therapeutic concept for patients with cancers bearing this KRAS alteration.

Up to 90% of patients with high-grade serous ovarian cancer (HGSC) will develop resistance to platinum-based chemotherapy, posing substantial therapeutic challenges due to a lack of universally druggable targets. Leveraging BenevolentAI's artificial intelligence (AI)-driven approach to target discovery, we screened potential AI-predicted therapeutic targets mapped to unapproved tool compounds in patient-derived 3D models. This identified TNIK, which is modulated by NCB-0846, as a novel target for platinum-resistant HGSC. Targeting by this compound demonstrated efficacy across both in vitro and ex vivo organoid platinum-resistant models. Additionally, NCB-0846 treatment effectively decreased Wnt activity, a known driver of platinum resistance; however, we found that these effects were not solely mediated by TNIK inhibition. Comprehensive AI, in silico, and in vitro analyses revealed CDK9 as another key target driving NCB-0846's efficacy. Interestingly, TNIK and CDK9 co-expression positively correlated, and chromosomal gains in both served as prognostic markers for poor patient outcomes. Combined knockdown of TNIK and CDK9 markedly diminished downstream Wnt targets and reduced chemotherapy-resistant cell viability. Furthermore, we identified CDK9 as a novel mediator of canonical Wnt activity, providing mechanistic insights into the combinatorial effects of TNIK and CDK9 inhibition and offering a new understanding of NCB-0846 and CDK9 inhibitor function. Our findings identified the TNIK-CDK9 axis as druggable targets mediating platinum resistance and cell viability in HGSC. With AI at the forefront of drug discovery, this work highlights how to ensure that AI findings are biologically relevant by combining compound screens with physiologically relevant models, thus supporting the identification and validation of potential drug targets.

Trophoblast cell surface antigen 2 (TROP2) exhibits aberrant expression in pancreatic cancer, correlating with metastasis, advanced tumor stage, and poor prognosis in patients with pancreatic ductal adenocarcinoma. TROP2 has been recognized as a promising therapeutic target for antibody-drug conjugates (ADC), as evidenced by the approval of the anti-TROP2 ADC Trodelvy for the treatment of triple-negative breast cancer (TNBC). In this study, we report the generation of novel second-generation amanitin-based ADCs (ATAC) targeting TROP2, comprising the humanized RS7 antibody of Trodelvy (hRS7) and the highly potent payload amanitin. The specific in vitro binding, efficient antigen internalization, and high cytotoxicity of hRS7 ATACs with EC50 values in the picomolar range in TROP2-expressing cells constituted the foundation for preclinical in vivo evaluation. The hRS7 ATACs demonstrated a significant reduction in tumor growth in vivo in subcutaneous xenograft mouse models of pancreatic cancer and TNBC at well-tolerated doses. The antitumor efficacy correlated with the level of TROP2 expression on the tumors and the in vivo tumor uptake of the ATACs. The long half-life of 9.7 to 10.7 days of hRS7 ATACs without premature payload release in serum supported a high therapeutic index. Notably, the efficacy of the hRS7 ATACs was superior to that of Trodelvy with complete tumor eradication in both refractory pancreatic cancer and TNBC xenograft models. In summary, hRS7 ATACs represent a highly effective and well-tolerated targeted therapy, and our data support their development for pancreatic cancer and other TROP2-expressing tumors.

Most of the triple-negative phenotypes or basal-like molecular subtypes of breast cancers are associated with aggressive clinical behavior and show poor disease prognosis. Current treatment options are constrained, emphasizing the need for novel combinatorial therapies for this particular tumor subtype. Our group has demonstrated that functionally active p21-activated kinase 1 (PAK1) exhibits significantly higher expression levels in clinical triple-negative breast cancer (TNBC) samples compared with other subtypes, as well as adjacent normal tissues. Low PAK1 expression in TNBC was significantly linked to better prognosis, with improved overall survival (P = 0.00236) and relapse-free survival (P = 0.0314), as shown by Gene expression-based Outcome for Breast cancer Online analysis. To confirm the role of PAK1 as a therapeutic target and to discover novel synergistic chemotherapy drug combinations, we conducted a drug combination screen using TNBC cell lines and a mouse metastatic tumor cell line. We identified the combined inhibition of PAK1 inhibitor NVS-PAK1 with doxorubicin/paclitaxel/methotrexate as a synergistic novel therapeutic approach for treating metastatic TNBC to improve overall survival. This study also indicated a reduction in the effective dosage of the chemotherapeutic drug when combined with NVS-PAK1. Our study demonstrates that combining NVS-PAK1 with each individual chemotherapeutic drug such as doxorubicin, paclitaxel, and methotrexate resulted in decreased colony formation, reduced wound-healing capability, and diminished migratory and invasive potential in both TNBC cell lines and 4T1 in vitro. These findings were further validated in orthotopic mouse mammary tumors, confirming that simultaneous PAK1 inhibition alongside chemotherapy significantly enhanced antitumor efficacy and reduced metastasis.

In patients with the rare adult-type granulosa cell tumor (aGCT), surgery is the primary treatment for both primary and recurrent disease. In cases of inoperable disease, systematic therapy is administered, but variable response rates and drug resistance complicate predicting the most effective therapy. Drug screen testing on patient-derived cell lines may offer a solution. In a national prospective study on aGCT, fresh tissue was cultured into 2D cell lines, testing 27 clinical and experimental drugs. Dose-response curves and synergy were calculated using GraphPad Prism and CompuSyn software. We established 34 patient-derived cell lines from tissue of 20 patients with aGCT. Of these, seven patients had a primary diagnosis of aGCT and 13 patients had recurrent disease. In eight patients, multiple tumor locations were cultured. On each cell line, 10 monotherapies and 17 combinations of drugs were tested. Carboplatin/gemcitabine showed efficacy and synergy in almost all patient-derived cell lines. Synergy could not be detected in the regular carboplatin/paclitaxel and carboplatin/etoposide combinations. Experimental combinations alpelisib/fulvestrant and alpelisib/gemcitabine showed efficacy of more than 75%. Drug screens on patient-derived tumor cell lines reflect the reality of the variable response of systemic therapy in patients with aGCT. In future research, this technique may be used to personalize the systemic treatment of patients with aGCT in a clinical study. The good response to carboplatin/gemcitabine in our patient-derived cell lines can then be confirmed in a clinical setting.

CD47 is a cell-surface glycoprotein that is expressed on normal human tissues and plays a key role as a marker of self. Tumor cells have co-opted CD47 overexpression to evade immune surveillance, and thus blockade of CD47 is a highly active area of clinical exploration in oncology. However, clinical development of CD47-targeted agents has been complicated by its robust expression in normal tissues and the toxicities that arise from blocking this inhibitory signal. Furthermore, pro-phagocytic signals are not uniformly expressed in tumors, and antibody blockade alone is often not sufficient to drive antitumor activity. The inclusion of an IgG1 antibody backbone into therapeutic design has been shown to not only serve as an additional pro-phagocytic signal but also exacerbate toxicities in normal tissues. Therefore, a need persists for more selective therapeutic modalities targeting CD47. To address these challenges, we developed SGN-CD47M, a humanized anti-CD47 IgG1 mAb linked to novel masking peptides through linkers designed to be cleaved by active proteases enriched in the tumor microenvironment (TME). Masking technology has the potential to increase the amount of drug that reaches the TME while concomitantly reducing systemic toxicities. We demonstrate that SGN-CD47M is well tolerated in cynomolgus monkeys and displays a 20-fold improvement in tolerability to hematologic toxicities when compared with the unmasked antibody. SGN-CD47M also displays preferential activation in the TME that leads to robust single-agent antitumor activity. For these reasons, SGN-CD47M may have enhanced antitumor activity and improved tolerability relative to existing therapies that target the CD47-signal regulatory protein α interaction.