STATEMENT REGARDING PRIOR DISCLOSURES BY THE INVENTOR OR A JOINT INVENTOR UNDER 37 C.F.R. 1.77(B)(6)

Part of the subject matter of the invention described in the present application was published by the inventors, Chen-Sheng Yeh, Liu-Chun Wang, Zheng-Zhe Chen, and Chia-Hao Su in an article titled “Low Dose of X-Ray-Excited Long-Lasting Luminescent Concave Nanocubes in Highly Passive Targeting Deep-Seated Hepatic Tumors;” this article was published online by Advanced Materials on 18 Oct. 2019. Therefore, the publications or disclosures was made by and/or originated from all member of the inventive entity of the present invention less than one year before the filing date of the present application. A copy of the article is provided in a concurrently filed Information Disclosure Statement pursuant to the guidance of 78 Fed. Reg. 11076 (Feb. 14, 2013).

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present disclosure relates to nanoprobes suitable for biological imaging. More particularly, the disclosure invention relates to modified chromium-doped zinc nanocubes, their production methods and uses thereof.

2. Description of Related Art

In vivo imaging faces two substantial challenges: the poor signal-to-noise (S/N) ratios due to tissue autofluorescence, and limited tissue penetration depth under in situ external excitation. To avoid tissue autofluorescence interference, several types of nanoprobes (e.g. Ag2S, carbon nanotubes) have been developed to emit near-infrared (NIR) wavelengths in the tissue transparency window for in vivo imaging to eliminate background noise. Alternatively, the long-lasting luminescence (LLL) nanophosphors characterized by luminescence persisting after excitation has ceased can improve the S/N signal by removing tissue background noise from in situ excitation. Usually, the lifetime of the biological luminescent signal would decay completely in nanoseconds, thus the LLL could be collected after the short-lived biological background. LLL nanophosphors are materials capable of storing radiation in the form of trapped electrons and holes, slowly emitting photons following carrier recombination.

Another concern in biological imaging using nanoprobes is the choice of the external excitation source to overcome the penetration limitation. Accordingly, significant work has focused on the fabrication of nanometric light-absorbing materials such as upconversion nanocrystals and semiconducting quantum dots which can be activated by the NIR-I, -II, and -III wavelengths. Recent studies using chromium-doped zinc gallate have demonstrated that the LLL of ZnGa2O4:Ce3+(ZGC) can be illuminated by red light. However, red light or even NIR source still suffers from tissue penetration limitations. For example, the first NIR (NIR-I) biological window has only 1 mm penetration and the 1,064 nm residing in the second NIR (NIR-II) tissue transparency region displays penetration of 5 mm. Accordingly, X-ray sources would be the optimal choice for imaging deeper tissue.

To eliminate tissue autofluorescence interference and overcome the tissue penetration limitation for in situ excitation, nanophosphors exhibiting LLL subjected to X-ray excitation show considerable promise for in vivo imaging. However, few bioimaging studies have used X-rays as an excitation source in LLL nanophosphors. Most previous work has on long-lasting luminescence mechanisms. More recently, activation of ZGC in red light has shifted from ex vivo excitation to in vivo excitation for LLL nanophosphors in bioimaging. ZGC exhibits a cubic spinel structure and is viewed as one of the most representative LLL phosphors, excitable by UV and visible sources. The radioluminescence of X-ray irradiation (immediately or delayed), has also been seen in ZGC powder- and nanosized-forms. Unfortunately, nanoscale ZGC typically exhibits an agglomerative or clustered morphology lacking well-defined cubic shape.

Accordingly, there exist in this art a need of improved ZGC nanoprobes with long lasting luminescence suitable for imaging deeper tissue.

SUMMARY

The following presents a simplified summary of the disclosure in order to provide a basic understanding to the reader. This summary is not an extensive overview of the disclosure and it does not identify key/critical elements of the present invention or delineate the scope of the present invention. Its sole purpose is to present some concepts disclosed herein in a simplified form as a prelude to the more detailed description that is presented later.

According to one aspect of the present disclosure, there is provided a modified chromium-dpoed zinc gallate (ZGC) nanocube, which is characterized in having a concave surface modified with (3-aminopropyl)triethoxysilane (APTES).

Additionally or optionally, each of the modified ZGC nanocube further comprises a plurality of polyethylene glycol (PEG) molecules independently linked to the APTES via an amide bond formed there between.

According to embodiments of the present disclosure, the modified ZGC nanocubes or the pegylated, modified nanocubes independently produce long lasting luminescence (LLL) that lasts for at least 1.5 hours after being excited with a dose of X-ray, preferably, the LLL lasts for at least 3 hours; more preferably, the LLL lasts for at least 5 hours.

According to the second aspect of the present disclosure, there is provided a method of producing a modified ZGC nanocube. The method includes steps of:(a) respectively reacting zinc nitrate and gallium nitrate with a base, thereby forming zinc hydroxide and gallium hydroxide;(b) mixing the zinc hydroxide and the gallium hydroxide respectively produced in the step (a) and chromium nitrate with water to give a first mixture;(c) adding a chelating agent and toluene to the first mixture to give a second mixture;(d) autoclaving the second mixture to produce a ZGC nanocube; and(e) silanizing the ZGC nanocube with (3-aminopropyl)triethoxysilane (APTES) to produce the modified ZGC nanocube.

Examples of the base suitable for use in the step (a) include but are not limited to, ammonium hydroxide, sodium hydroxide, potassium hydroxide and the like.

The chelating agent suitable for use in the step (c) may be C16-20fatty acid, carboxylated PEG or ascorbic acid. Suitable examples of C16-20fatty acid include, but are not limited to, palmitic acid, oleic acid, margaric acid, stearic acid, nonadecanoic acid, eicosanoic acid, palmitoeic acid, elaidic acid, vaccenic acid, linoleic acid, linolelacidic acid, α-linolenic acid, stearidonic acid, paullinic acid, gondoic acid, mead acid, dihomo-γ-linolenic acid, and arachidonic acid.

According to embodiments of the present disclosure, in the step (d), the autoclave is conducted at a temperature of 220° C. for 3 days; and in the step (e), the silanization is conducted at a temperature of 60° C. for 18 hours.

According to optional embodiments of the present disclosure, the method may further include a step of pegylating the modified ZGC nanocube by reacting with amine-PEG-acid in the presence of 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS), thereby producing a pegylated modified ZGC nanocube, in which the amine-PEG-acid has a molecular weight of about 3,400.

According to preferred embodiments of the present disclosure, the modified ZGC nanocube and the pegylated modified ZGC nanocube respectively produce long lasting luminescence (LLL) that lasts for at least 1.5 hours after being excited with X-ray; preferably, the LLL lasts for at least 3 hours after being excited with X-ray; more preferably, the LLL lasts for at least 5 hours after being excited with X-ray.

According to the second aspect of the present disclosure, there is provided a method of imaging an area of interest (AOI) in a subject. The method includes steps of:(a) administering a sufficient amount of the pegylated modified ZGC nanocube of the present disclosure to the AOI; and(b) irradiating the subject with a dose of X ray thereby producing the image of the AOI.

According to embodiments of the present disclosure, in the step (b), the dose of X ray is no more than 3 Gy. Preferably, the dose of X ray is about 0.5 Gy.

According to embodiments of the present disclosure, the AOI is a cancer.

Examples of cancer suitable for imaging by the present method include, but are not limited to, bone cancer, brain cancer, breast cancer, colon cancer, cervical cancer, Ewing's sarcoma, esophageal cancer, hepatic cancer, head and neck cancer, larynx cancer, melanoma, multiple myeloma, nasopharynx cancer, non-small-cell lung cancer, non-melanoma skin cancer, neuroblastoma, pancreatic cancer, prostate cancer, retinoblastoma, rectal cancer, small-cell lung cancer, testicular cancer, thyroid cancer, and Wilms' tumor.

According to one preferred embodiment of the present disclosure, the cancer is hepatic cancer.

According to embodiments of the present disclosure, the subject is a mammal. Preferably, the subject is a human.

Many of the attendant features and advantages of the present disclosure will becomes better understood with reference to the following detailed description considered in connection with the accompanying drawings.

