Research Areas

Summary of Nanodrugs and Nanoimaging Agents

Nano bioconjugates were engineered based on a natural biodegradable nanoplatform. The drug/imaging agents are specifically delivered crossing multiple bio-barriers such as the BBB and the cancer cell membrane to deliver the anti-cancer drugs to the cytoplasm/nuclei of the tumor cells using one or several targeting monoclonal antibodies or peptides. This controllable unique drug releases tumor growth inhibiting agents specifically into cancer cells without affecting normal surrounding cells. Minimal toxicity was demonstrated for nanodrug tumor delivery for temozolomide, platinum drugs and doxorubicin.

MRI Virtual Biopsy for Brain Lesions Is Used When it Is Impossible to Obtain Brain Tissue for Diagnosis

research, HER2, metastatic breast tumor, Ljubimova Laboratory, Julia Ljubimova, EGFR+ glioblastoma multiforme, Cedars-Sinai

Figure 1. Detection of HER2+ metastatic breast tumor in the presence of EGFR+ glioblastoma multiforme (GBM) (double tumor model) using HER2-targeting NIA. (Right) MRI scans of mice with double tumors, a primary GBM (U87MG, EGFR+) in the left hemisphere and metastatic breast cancer (BT-474, HER2+) in the right hemisphere after IV injection of targeted NIA (P/Gd-DOTA/trastuzumab/MsTfR-mAb/Alexa-680). Both tumors showed contrast at 20 minutes. At later times, only HER2+ tumor retained high contrast due to specific accumulation of targeted agent, whereas little contrast was seen in the EGFR+ non-targeted tumor. From Patil R et al. ACS Nano. 2015;5:5594-5608.

Discovery of the New Cancer Biomarker, Laminin 411

Institute researchers used leading-edge gene-sequencing analysis to discover a new cancer biomarker: laminin-411. Human gliomas and invasive breast cancer excessively produce laminin-411, which plays an important role in the ability of tumor cells to spread and grow (Cancer Res. 2001;61:5601-5610. Breast Cancer Res. 2005;7:411-421.). This biomarker was later analyzed in a number of human gliomas and showed a significant correlation with glial tumor grade, time to recurrence and patients' survival times (Cancer. 2004;101:604-612.). Inhibition of this marker and drug delivery into the brain in vivo was developed, and glioma tumor reduction was achieved (PNAS USA. 2010;107:18143-18148.). This test is being used to evaluate the biological behavior of gliomas in order to better plan individualized therapeutic treatment and follow-up regimens for each patient. An ongoing clinical trial is underway in the Neurosurgery Department and Department of Pathology and Laboratory Medicine at Cedars-Sinai for laminin-411 as a glial tumor biomarker (about 400 patients have been evaluated).

The Ljubimova Laboratory has also broken new ground, the team has developed nanomedicine technology to inhibit the synthesis of cancer-specific extracellular matrix protein, laminin-411, and to engineer a novel drug delivery system to reduce gliomas in the brain in vivo

inhibition of Laminin 411, Notch Pathway, infographic, diagram, Ljubimova Laboratory, Julia Ljubimova, EGFR+ glioblastoma multiforme, Cedars-Sinai

Figure 2. Scheme of Inhibition of Laminin 411-Notch Pathway. Integrin b1, Dll4, Jagged1 and Notch1; and cancer stem cells markers: Notch-3;CD133; Nestin; and c-Myc in mice after treatment with nanoconjugate. The expression of Nestin and c-Myc was found close to tumor blood vessels positive for laminin β1 staining. (From Tao S, et al. Cancer Res. 2019;79:1239-1251.)

In a panel of 226 patient brain glioma samples, the Ljubimova Lab found a clinical correlation between the expression of tumor vascular laminin-411 (α4β1γ1) with higher tumor grade and with expression of cancer stem cell (CSC) markers including Notch pathway members, CD133, Nestin and c-Myc. Laminin-411 overexpression also correlated with higher recurrence rate and shorter survival of GBM patients. The lab also showed that depletion of laminin-411 α4 and β1 chains with CRISPR/Cas9 in human GBM cells led to reduced growth of resultant intracranial tumors in mice, and significantly increased survival of host animals compared to mice with untreated cells. Inhibition of laminin-411 suppressed Notch pathway in normal and malignant human brain cell types. A nano bioconjugate potentially suitable for clinical use and capable of crossing BBB was designed to block laminin-411 expression. Nano bioconjugate treatment of mice carrying intracranial GBM significantly increased animal survival and inhibited multiple CSC markers including the Notch axis. This study describes an efficient strategy for GBM treatment via targeting a critical component of the tumor microenvironment largely independent of heterogeneous genetic mutations in glioblastoma.

Extracellular Matrix, GBM Development, infographic, diagram, Ljubimova Laboratory, Julia Ljubimova, Cedars-Sinai

Figure 3. Extracellular Matrix Contributes to GBM Development. As laminin-411 is overexpressed during glioma progression, the lab has tested if its loss in GBM would impair tumor growth. The lab disrupted laminin-411 α4 and β1 genes in two established human GBM cell lines and two patient-derived primary GBM lines TS543 and TS576 (not shown) using CRISPR/Cas9 (left A) and confirmed specific elimination of laminin a4 and b1 chains. The lab then inoculated U87MG and LN229 GBM cells with laminin α4 and β1 double knockout (DKO) along with their wild type (WT) counterparts intracranially into nude mice (n=8 in WT groups and n=10 in DKO groups (left B). After four weeks, tumor sizes were measured by MRI. Compared to WT tumors, DKO tumors were significantly smaller (left C). Consistent with imaging results, animals carrying U87MG (moderately invasive) or LN229 (highly invasive) laminin DKO tumors had significantly longer survival than animals with WT tumors (left D and E). After the BBB crossing, polymeric nano bioconjugate released molecular inhibitors into the cytoplasm of glioma cells in vivo (verified by confocal microscopy) preventing the synthesis of laminin-411 with decreased tumor size by 90% (right A-C) and significant increased survival. From Cancer Res. 2019;79:1239-1251. 

Immunotherapy is one of the fastest developing approaches in clinical oncology with successful treatment of different cancers. However, the unique immune environment of the central nervous system needs consideration when pursuing immunotherapeutic approaches for gliomas. Treatment options are limited, in part because of inefficient drug delivery across the BBB.

The Ljubimova Laboratory developed new nano immunoconjugates (NICs) based on natural biopolymer scaffold, poly(-L-malic acid), with covalently attached a-CTLA-4 and/or a-PD-1 for delivery across the BBB and activation of local brain anti-tumor immune response. NIC treatment of mice bearing intracranial GL261 GBM resulted in an increase of anti-tumor immune cells such as CD8+ T cells, IFN+ natural killers and natural killer T cells and macrophages with a decrease of regulatory T cells (Tregs) in the brain tumor area. Survival of GBM-bearing mice treated with combination of NICs was significantly longer compared to animals treated by single checkpoint inhibitor-bearing NICs or free a-CTLA-4 and a-PD-1. Our study demonstrates trans-BBB delivery of nanopolymer-conjugated checkpoint inhibitors as an effective treatment of GBM via activation of both systemic and local privileged brain tumor immune response.

Mechanism, Local Brain, Immune System Activation, infographic, diagram, Ljubimova Laboratory, Julia Ljubimova, Cedars-Sinai

Figure 4. Proposed Mechanism of the Local Brain Immune System Activation After Nano Immmunoconjugates (NICs) Treatment. The proposed mechanism of synergistic treatment with a-CTLA-4 and a-PD-1 mAbs when they cross BBB as part of a nano immunodrug. (1) a-PD-1 initiates central pathway inhibition with cancer cell attack; (2) a-PD-1 activates cytotoxic T lymphocytes and (3) a-CTLA-4 provides local inhibition of Treg and activation of natural killers and macrophages. (From Galstyan A, et al. Nat Commun. 2019.)

Crosstalk between CK2 and EGFR/EGFRvIII pathways was found by blocking two glioma specific markers. The active drug component blocks several cancer-specific tumor markers at the same time for multiple tumors: EGFR/EGFRvIII-wild/mutated epidermal growth factor receptor, CK2, a master signaling regulator serine-threonine protein kinase, and HER2/neu. A significant survival increase has been shown in tumor-bearing animals (preclinical data) after drug administration. Using this technology, it is possible to block a combination of several unique markers for each patient at the same time, providing a synergistic effect. (From J Cont Release. 2016;28(244):14-23.)

Mini Nanodrugs: A Powerful Improvement in Synthesis and Function of Nanomedicines

Nanomedicine, February 2017, cover, Ljubimova Laboratory, Julia Ljubimova, Cedars-Sinai

Figure 5. Mini drug inhibition of HER2-positive breast cancer model om mic. (From Ding H, et al. Nanomedicine. 2017;13:631-639.) 

Introduction of Mini Platforms for Delivery Through BBB

trileucine, angiopep-2, polyanionic polymalic acid, nanodrugm blood brain barrier, research, Ljubimova Laboratory, Julia Ljubimova, Cedars-Sinai

Figure 6. A combination of trileucine and angiopep-2 drives a polyanionic polymalic acid nanodrug across BBB. A possible boosting mechanism is depicted functioning in productive positioning of shuttle peptides for receptor binding BBB penetration is programmed by the composition of the nanoconjugate (sizes 5-7 nm). (From Israel, et al. ACS Nano. 2019;13:1253-1271.)

Mini Nano Agents Can Be Used as Tumor "Paints"

Nano Imaging, research, infographic, diagram, Ljubimova Laboratory, Julia Ljubimova, Cedars-Sinai

Figure 7. Left A-C: Essential features of a Nano Imaging. Imaging agents are clearanceing fast from the blood (A) to achieve prolonged retention in the tumor with a very low fluorescence background (B). Distribution during early agent clearing from the blood (C). Right A-D: Assessment of the resection precision after H&E staining (A, B) indicating pieces of tumor and also of falsly resected healthy brain. Comparison and statistics of resection analysis (C, D) comparing resection under NIR and under (normal) white light illumination. (From Patil R,  et al, Biomaterials. 2019;206:146-159.)

Contact the Ljubimova Lab

127 S. San Vicente Blvd.
Advanced Health Sciences Pavilion, Eighth Floor
Los Angeles, CA 90048

Helena Kozlova, MBA