by Xiaoyue Chen, 2008-8-13

• Background and Goals

It is generally accepted idea that the future treatment for cancer relies on early detection of cancer lesions, as well as efficient and specific delivery of drugs to the cancer cell target. The detection of stage 1 cancer is associated with a > 90% 5-year survival rate [1] while conventional anatomic imaging typically cannot detect cancers until they reach > 1cm diameter [2]. Recently, molecular imaging especially with quantum dots (QDs) covalently linked to biorecognition molecules such as peptides, antibodies, nucleic acids or small-molecule ligands is expected to play an important role in future cancer diagnosis. In comparison with organic fluorophores, QDs have many unique optical properties including size-tunable emission over a broad wavelength, large absorption coefficient across a wide spectral range, high photostability and quantum yield (Figure 1) [3-4] .

At present, however, little is known about these bio-functionalized QDs, especially the choosing of targeting molecules, conjugation techniques, and the metabolism and distribution of QDs injected into living animals. The variability among targeting molecules, as well as conjugation chemistry of many molecular targeting platforms have not been well-characterized in vivo yet. Since there are so many variances of cancer types, it will be very meaningful to get the insight into the QDs modified by other targeting molecules and conjugation methods.

In recent years, many new molecular approaches to cancer diagnosis and treatment are being investigated. One of them is T cell-based strategy targeting tumor antigen, like p53, which is overexpressed and presented on HLA class I molecules as peptides in many types of tumors. Soluble single-chain TCR (scTCR) recognizing wild-type human p53 peptide (aa 264-272) has been constructed to detect tumor cells displaying as few as 500 peptide/MHC complexes in flow cytometry [5]. Another strategy is proteolytic activated peptides-based, without requiring specific receptors. Cell-penetrating peptides (CPPs) activated by matrix metalloproteinases (MMPs), which are overexpressed in tumor invasion and metastasis, have already been devised for the internalization of reporters into tumor cells [6]. It will be interesting to functionalize QDs with some new targeting molecules like scTCR and CPPs, in which a wider spectrum of diagnostic QDs can be developed while more conjugation varieties can be tested.

What is more, the extraordinary optical properties of QDs can also revolutionize some current cancer diagnosis techniques which are radioactivity-involved. Diagnosis and treatment of thyroid cancer based on thyroid cells uptaking radioactive iodine is one of them. Considering the well-known drawback of exposure to radioactivity, we believe QDs functionalized with molecules targeting thyroid cells like antibody against thyroid stimulating hormone receptor (TSHR) and TSH analogs will be a much safer and promising diagnostic method.

The overall objective of this project is developing a novel class of diagnostic and therapeutic agent with high selectivity of tumor. We are now working on developing QDs functionalized by different cancer targeting molecules including scTCR, MMP-activatable peptide (MAP), anti-TSHR antibody, etc (Figure 2). We aim to discovering more knowledge about bio-conjugation of nanoparticles and developing nanoparticles for biomedical applications.

• Methods

Successful bioconjugation depends on the prudent choice of crosslinkers and reaction conditions. We are now working on several different kinds of crosslinkers including homobifunctional NHS/EDC and heterobifunctional SMCC (Figure 3).

FPLC gel filtration and CsCl gradient ultracentrifuge are used for the purification of desired conjugates (Figure 4).

Cell binding and internalization experiments are facilitated by flow cytometry and fluorescence microscopy (Figure 4).

We can also achieve other facilities for characterization of the size, surface charge of QD conjugates here in Cornell.

• Future Plans

Besides bioconjugation of QDs, our lab is also working on some other nanoparticles for biomedical use including liposomes, SPIO and C-dots. This offers great opportunity for the comparison and learning on the properties and functions of different imaging agents. On the other hand, QDs has been reported to have great potential for applications in photodynamic therapy due to its energy transfer effect (7-8). Therefore future development of tumor targeting QDs from diagnosis to therapy is very promising.

• References

1. R. Etzioni, et al. Early detection: The case for early detection. Nat. Rev. Cancer 3, 243 (2003).

2. Weissleder R. Molecular imaging in cancer. Science 312, 1168-1171 (2006).

3. Bruchez M Jr, Moronne M, Gin P, Weiss S & Alivisatos AP. Semiconductor nanocrystals as fluorescent biological labels. Science 281, 2013-2015 (1998).

4. Rosenthal SJ, et al. Targeting cell surface receptors with ligand-conjugated nanocrystals. J. Am. Chem. Soc.124, 4586-4594 (2002).

5. Zhu X, et al. Visualization of p53(264-272)/HLA-A*0201 complexes naturally presented on tumor cell surface by a multimeric soluble single-chain T cell receptor. J. Immunol. 176(5), 3223-3232 (2006)

6. Jiang T, et al. Tumor imaging by means of proteolytic activation of cell-penetrating peptides. Proc. Natl. Acad. Sci. 101(51), 17867-17872 (2004)

7. Bakalova R, et al. Quantum dot anti-CD conjugates: are they potential photosensitizers or potentiators of classical photosensitizing agents in photodynamic therapy of cancer? Nano Letters 4(9), 1567-1573 (2004)

8. Samia AC, Dayal S, Burda C. Quantum dot-based energy transfer: perspectives and potential for applications in photodynamic therapy. Photochem. Photobiol. 82(3), 617-625 (2006)

• Related Links

Collaborators

Ocean Nanotech http://www.oceannanotech.com/home.html

Altor Bioscience http://www.altorbioscience.com/

Trophogen http://www.trophogen.com/

Learn more about the studies on QDs at:

Quantum dots on Wiki http://en.wikipedia.org/wiki/Quantum_dot

Evident Technologies http://www.evidenttech.com/