Strategies To Inhibit Tumor Associated Integrin Receptors: Rationale for Dual and Multi-Antagonists

Sheldrake, Helen M.; Patterson, Laurence H.

doi:10.1021/jm5000547

Cited by 80 publications

(108 citation statements)

References 146 publications

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“…EETI 2.5F is unique in its ability to target a broad spectrum of tumor-associated integrins, a feature that has not been achieved with any antibody to date, which instead only bind av-containing integrins or a5b1 integrin. Targeting multiple integrin receptors is potentially beneficial for reducing drug resistance and tumor growth, as cancer cells can change their integrin repertoire in response to drug treatment (36). Integrins, particularly a6b4, a6b1, avb5, a2b1, and a3b1 (21), are found to be present on epithelial tissues and are also upregulated in wound healing.…”

Section: Discussionmentioning

confidence: 99%

Targeted Drug Delivery with an Integrin-Binding Knottin–Fc–MMAF Conjugate Produced by Cell-Free Protein Synthesis

Currier

Ackerman

Kintzing

et al. 2016

Molecular Cancer Therapeutics

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Antibody-drug conjugates (ADC) have generated significant interest as targeted therapeutics for cancer treatment, demonstrating improved clinical efficacy and safety compared with systemic chemotherapy. To extend this concept to other tumor-targeting proteins, we conjugated the tubulin inhibitor monomethyl-auristatin-F (MMAF) to 2.5F-Fc, a fusion protein composed of a human Fc domain and a cystine knot (knottin) miniprotein engineered to bind with high affinity to tumorassociated integrin receptors. The broad expression of integrins (including avb3, avb5, and a5b1) on tumor cells and their vasculature makes 2.5F-Fc an attractive tumor-targeting protein for drug delivery. We show that 2.5F-Fc can be expressed by cellfree protein synthesis, during which a non-natural amino acid was introduced into the Fc domain and subsequently used for site-specific conjugation of MMAF through a noncleavable linker. The resulting knottin-Fc-drug conjugate (KFDC), termed 2.5F-Fc-MMAF, had approximately 2 drugs attached per KFDC. 2.5F-Fc-MMAF inhibited proliferation in human glioblastoma (U87MG), ovarian (A2780), and breast (MB-468) cancer cells to a greater extent than 2.5F-Fc or MMAF alone or added in combination. As a single agent, 2.5F-Fc-MMAF was effective at inducing regression and prolonged survival in U87MG tumor xenograft models when administered at 10 mg/kg two times per week. In comparison, tumors treated with 2.5F-Fc or MMAF were nonresponsive, and treatment with a nontargeted control, CTRL-Fc-MMAF, showed a modest but not significant therapeutic effect. These studies provide proofof-concept for further development of KFDCs as alternatives to ADCs for tumor targeting and drug delivery applications.

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Section: Discussionmentioning

confidence: 99%

Targeted Drug Delivery with an Integrin-Binding Knottin–Fc–MMAF Conjugate Produced by Cell-Free Protein Synthesis

Currier

Ackerman

Kintzing

et al. 2016

Molecular Cancer Therapeutics

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“…Since the cyclic RGD peptides, such as c(RGDfK), are antagonists for α v β 3 and α v β 5 integrins, 4,36−39,62 it is reasonable to believe that bioconjugates FITC-RGD 2 , FITC-3P-RGD 2 , and FITC-Galacto-RGD 2 will target both integrin α v β 3 and integrin α v β 5 . We were interested in dimeric cyclic RGD peptides because they have significantly higher integrin α v β 3 /α v β 5 binding affinity than their monomeric analogs.…”

Section: Introductionmentioning

confidence: 99%

FITC-Conjugated Cyclic RGD Peptides as Fluorescent Probes for Staining Integrin α_vβ₃/α_vβ₅ in Tumor Tissues

Zheng

Czerwiński

et al. 2014

Bioconjugate Chem.

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This study sought to evaluate FITC-conjugated cyclic RGD peptides (FITC-RGD2, FITC-3P-RGD2, and FITC-Galacto-RGD2) as fluorescent probes for in vitro assays of integrin αvβ3/αvβ5 expression in tumor tissues. FITC-RGD2, FITC-3P-RGD2, and FITC-Galacto-RGD2 were prepared, and their integrin αvβ3/αvβ5 binding affinity was determined using the displacement assay against 125I-echistatin bound to U87MG glioma cells. IC50 values of FITC-Galacto-RGD2, FITC-3P-RGD2, and FITC-RGD2 were calculated to be 28 ± 8, 32 ± 7, and 89 ± 17 nM, respectively. The integrin αvβ3/αvβ5 binding affinity followed a general trend: FITC-Galacto-RGD2 ∼ FITC-3P-RGD2 > FITC-RGD2. The xenografted tumor-bearing models were established by subcutaneous injection of 5 × 106 tumor cells into shoulder flank (U87MG, A549, HT29, and PC-3) or mammary fat pad (MDA-MB-435) of each athymic nude mouse. Three to six weeks after inoculation, the tumor size was 0.1–0.3 g. Tumors were harvested for integrin αvβ3/αvβ5 staining, as well as hematoxylin and eosin (H&E) staining. Six human carcinoma tissues (colon cancer, pancreatic cancer, lung adenocarcinoma, squamous cell lung cancer, gastric cancer, and esophageal cancer) were obtained from recently diagnosed cancer patients. Human carcinoma slides were deparaffinized in xylene, rehydrated with ethanol, and then used for integrin αvβ3/αvβ5 staining, as well as H&E staining. It was found that the tumor staining procedures with FITC-conjugated cyclic RGD peptides were much simpler than those with the fluorescence-labeled integrin αvβ3 antibodies. Since FITC-RGD2, FITC-3P-RGD2, and FITC-Galacto-RGD2 were able to co-localize with the fluorescence-labeled integrin β3 antibody, their tumor localization and tumor cell binding are integrin αvβ3-specific. Quantification of the fluorescent intensity in five xenografted tumors (U87MG, MDA-MB-435, A549, HT29, and PC-3) and six human carcinoma tissues revealed an excellent linear relationship between the relative integrin αvβ3/αvβ5 expression levels determined with FITC-Galacto-RGD2 and those obtained with the fluorescence-labeled anti-human integrin β3 antibody. There was also an excellent linear relationship between the tumor uptake (%ID/g) of 99mTc-3P-RGD2 (an integrin αvβ3/αvβ5-targeted radiotracer) and the relative integrin αvβ3/αvβ5 expression levels from the quantification of fluorescent intensity in the tumor tissues stained with FITC-Galacto-RGD2. These results suggest that FITC-conjugated cyclic RGD peptides might be useful to correlate the in vitro findings with the in vivo imaging data from an integrin αvβ3/αvβ5-targeted radiotracer. The results from this study clearly showed that the FITC-conjugated cyclic RGD peptides (particularly FITC-3P-RGD2 and FITC-Galacto-RGD2) are useful fluorescent probes for assaying relative integrin αvβ3/αvβ5 expression levels in tumor tissues.

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“…Recent findings have shown that cancer cells change their integrin repertoire in response to drug treatment (Sheldrake and Patterson, 2014). Given the link between integrin expression and CYP3A4 activity illustrated here, integrin receptors may play a notable role in the development of drug resistance.…”

Section: Discussionmentioning

confidence: 77%

“…The diversity in these processes suggests that pathology of cancer as well as other noninfectious diseases in which integrins play a role (e.g., cardiovascular disease, diabetes, and wound healing) may not be attributed to a single integrin receptor, but instead are the end result of crosstalk between several integrin receptors. Thus, drugs targeting single as well as multiple integrin receptors have been developed for treatment of cancer and other diseases (Desgrosellier and Cheresh, 2010;Sheldrake and Patterson, 2014). In this context, data generated in our studies suggest that hepatic drug metabolism patterns can be significantly altered in response to these medications, which will most likely be given with other chemotherapeutic agents that are metabolized by CYP3A4, resulting in either an enhanced therapeutic effect or severe toxicities due to accumulation of unmetabolized by-products over time.…”

Section: Discussionmentioning

confidence: 91%

Integrin Receptors Play a Key Role in the Regulation of Hepatic CYP3A

Jonsson-Schmunk

Wonganan

Choi

et al. 2016

Drug Metabolism and Disposition

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Landmark studies describing the effect of microbial infection on the expression and activity of hepatic CYP3A used bacterial lipopolysaccharide as a model antigen. Our efforts to determine whether these findings were translatable to viral infections led us to observations suggesting that engagement of integrin receptors is key in the initiation of processes responsible for changes in hepatic CYP3A4 during infection and inflammation. Studies outlined in this article were designed to evaluate whether engagement of integrins, receptors commonly used by a variety of microbes to enter cellular targets, is vital in the regulation of CYP3A in the presence and absence of virus infection. Mice infected with a recombinant adenovirus (AdlacZ) experienced a 70% reduction in hepatic CYP3A catalytic activity. Infection with a mutant virus with integrin-binding arginine-glycine-aspartic acid (RGD) sequences deleted from the penton base protein of the virus capsid (AdDRGD) did not alter CYP3A activity. CYP3A mRNA and protein levels in AdlacZtreated animals were also suppressed, whereas those of mice given AdDRGD were not significantly different from uninfected control mice. Silencing of the integrin b-subunit reverted adenovirus-mediated CYP3A4 suppression in vitro. Silencing of the a-subunit did not. Suppression of integrin subunits had a profound effect on nuclear receptors pregnane X receptor and constitutive androstane receptor, whereas retinoid X receptor a was largely unaffected. To our knowledge, this is the first time that extracellular receptors, like integrins, have been indicated in the regulation of CYP3A. This finding has several implications owing to the important role of integrins in normal physiologic process and in many disease states.

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Strategies To Inhibit Tumor Associated Integrin Receptors: Rationale for Dual and Multi-Antagonists

Cited by 80 publications

References 146 publications

Targeted Drug Delivery with an Integrin-Binding Knottin–Fc–MMAF Conjugate Produced by Cell-Free Protein Synthesis

Targeted Drug Delivery with an Integrin-Binding Knottin–Fc–MMAF Conjugate Produced by Cell-Free Protein Synthesis

FITC-Conjugated Cyclic RGD Peptides as Fluorescent Probes for Staining Integrin α_vβ₃/α_vβ₅ in Tumor Tissues

Integrin Receptors Play a Key Role in the Regulation of Hepatic CYP3A

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Strategies To Inhibit Tumor Associated Integrin Receptors: Rationale for Dual and Multi-Antagonists

Cited by 80 publications

References 146 publications

Targeted Drug Delivery with an Integrin-Binding Knottin–Fc–MMAF Conjugate Produced by Cell-Free Protein Synthesis

Targeted Drug Delivery with an Integrin-Binding Knottin–Fc–MMAF Conjugate Produced by Cell-Free Protein Synthesis

FITC-Conjugated Cyclic RGD Peptides as Fluorescent Probes for Staining Integrin αvβ3/αvβ5 in Tumor Tissues

Integrin Receptors Play a Key Role in the Regulation of Hepatic CYP3A

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FITC-Conjugated Cyclic RGD Peptides as Fluorescent Probes for Staining Integrin α_vβ₃/α_vβ₅ in Tumor Tissues