When Viruses and Carbon Nanotubes Collide: A Novel Approach to Cancer Imaging

One of the biggest challenges in cancer treatment is finding and treating metastases, or small tumors spread throughout the body. It is best to identify and remove tumors before the process of metastasis begins; unfortunately, this can be difficult in some cancers because the symptoms patients present may not be immediately obvious as signs of cancer, and metastases are often undetectable using our current diagnostic methods. For example, ovarian cancer is often called the “silent killer” because many of the symptoms women notice first, such as abdominal pain or bloating, feeling full sooner than expected, or having difficulty urinating, are easily confused with other less serious causes [1]. Additionally, the ovaries are located deep inside the abdomen, so it is often difficult, if not impossible, for a doctor to feel any tumors during a routine pelvic exam [1,2]. Thus, it is not surprising that 80% of ovarian cancer cases are not detected until the cancer has progressed to more advanced stages and spread to other parts of the body [3].

 

The current method to remove ovarian cancer is a combination of surgery and chemotherapy [2]. During surgery, the doctor will visually inspect and remove both the tumor and some healthy tissue surrounding the tumor in case there are any residual cancer cells nearby that are invisible to the naked eye. However, this visual inspection method will miss sub-millimeter tumors that are located under deeper layers of tissue. By not removing these types of tumors, the cancer is very likely to return.

 

One potential solution is using fluorescent imaging as a portable and non-invasive way to observe cancer cells. Fluorescence imaging works by shining light of a certain wavelength on a material, which absorbs the energy and emits a different wavelength of light to be detected and recorded. If such a material could be attached to cancer cells, the presence, location and size of malignant tumors could be easily determined. It is with this idea in mind that scientists at the Massachusetts Institute of Technology (MIT) designed an imaging tool to probe for ovarian cancer cells.

Graphic by Stephanie DeMarco
Box 1: The electromagnetic spectrum is a representation of the different wavelengths of light. It is depicted here showing light with the longest wavelength on the left to light with the shortest wavelength on the right. NIR1 and NIR2 both represent specific ranges of wavelengths in the infrared part of the spectrum, with NIR1 corresponding to wavelengths from 650-900 nm and NIR2 corresponding to wavelengths from 950-1400 nm.

In innovative new work, researchers in the Bhatia and Belcher laboratories at MIT have developed a new tool to screen for small tumors that are embedded deep within tissues by combining a virus, carbon nanotubes, and a small molecule that specifically recognizes ovarian cancer cells [4]. To provide the structural support for the imaging probe, they started with M13 bacteriophage virus. M13 is a relatively simple virus that only infects bacteria. The researchers took advantage of M13’s rod-shaped structure by using it as a platform, as one would use a long and thin LEGO piece as the base to build a bigger structure.

 

Next, a fluorescent material was needed. Some imaging probes use a material that releases a wavelength of light in the near infrared first window (NIR1) part of the electromagnetic spectrum of light (Box 1). Light in the NIR1 wavelength has a few drawbacks for identifying tumors. Many materials that emit light in this wavelength lose the ability to continue to fluoresce after a certain period of time, a phenomenon called photobleaching [4]. It is important that the imaging probe not photobleach in order to give the surgeon enough time to find and remove the tumors. In addition, imaging probes that emit light in the NIR1 part of the spectrum cause the healthy tissue surrounding the cancer cells to appear to fluoresce (auto-fluorescence), which makes it impossible to differentiate between the boundaries of the tumor and the surrounding healthy tissue [5]. This is akin to trying to focus on the bright headlights of an oncoming car when driving at night. You know that the headlights is there, but you can’t make out where they are relative to the license plate because the entire front of the car looks bright.

Carbon nanotubes. Image credit: Michael Ströck, Licensed under the Creative Commons Attribution-Share Alike 3.0 Unported license.

In an effort to overcome these problems, the researchers took advantage of the fluorescent properties of carbon nanotubes. Carbon nanotubes are single atom-thick tubes made of carbon atoms arranged in a honeycomb structure. Just as diamond and graphite are made up of carbon atoms yet have strikingly different physical characteristics, the honeycomb arrangement of the carbon atoms in a nanotube give it unique properties, such as the ability to withstand extremely high pressures, conduct electricity, and when excited with light, to release light in the near infrared second window (NIR2) part of the electromagnetic spectrum [6,7]. Carbon nanotubes are less likely to photobleach than materials that fluoresce in NIR1 wavelengths, and NIR2 light causes less autofluorescence than NIR1 emitted light, which makes carbon nanotubes an excellent way to overcome the problems plaguing the previous imaging probes [5]. By engineering M13 to express a molecule that binds to carbon nanotubes, the researchers added a fluorescent component to their imaging probe. 

 

The final requirement was to target the probe to ovarian cancer cells. To do this, they took advantage of the fact that ovarian cancers have a distinguishing marker called SPARC (Secreted Protein, Acidic and Rich in Cysteines) [8]. The researchers modified M13 to include a small molecule that binds specifically to SPARC on ovarian cancer cells. As the complete imaging probe, M13 is able to bring the fluorescent carbon nanotubes specifically to ovarian cancer cells, so when light is shined on the abdomen either during a non-invasive diagnostic test or during surgery, the doctor will be able to see exactly where the tumors are located relative to the patient’s healthy tissue.

Figure 1: The researchers found that the probe, referred to as SBP-M13-SWNTs, did not photobleach and had a higher signal-to-noise ratio than the other imaging probes.

Figure 1: The researchers found that the probe, referred to as SBP-M13-SWNTs, did not photobleach and had a higher signal-to-noise ratio than the other imaging probes.

 

The researchers began by asking how well the probe performed compared to standard fluorescent markers. They found that the M13 probe did not photobleach even when continuously exposed to light for 30 minutes (Figure 1). In comparison, a different fluorescent marker that has been used to image ovarian cancer cells in humans, called FITC, did photobleach during the same 30 minute time period. They also found that the ratio of fluorescence of tumor-to-background (the signal-to-noise ratio) was 28-fold higher with the M13 probe than the previously used FITC marker, meaning that tumors were much more easily distinguished from the background healthy tissue using the carbon-nanotube device.

 

To see if the probe could also help improve tumor identification during surgery, the researchers first performed surgery on mice using only visual inspection, as is the current standard surgical procedure for ovarian cancer removal. After the initial surgery, they imaged using the fluorescent M13 probe and found that many small tumors and tumors located deep inside tissues were still present, which they had missed by looking with the naked eye (Figure 2). After the probe enabled the surgeons to see these tumors, they were able to remove almost all of these additional tumors from the mice.

This new probe has incredible implications for non-invasive ovarian cancer screening and surgical procedures, and it makes vast improvements on other fluorescent imaging tools currently available. Due to the building block-like nature of the device, it is likely that by swapping out the targeting molecule that directs the device specifically to ovarian cancer, it will be possible to target this device to other types of cancer cells. Thus, this technology has the potential to not only improve cancer screening for ovarian cancer patients but for patients with other cancers as well.

Figure 2: Using the probe to image during surgery, the researchers found more small and deeply embedded tumors. The white arrow points to a tumor that was only found and removed after imaging using the probe.

Figure 2: Using the probe to image during surgery, the researchers found more small and deeply embedded tumors. The white arrow points to a tumor that was only found and removed after imaging using the probe.


To find out more information about the scientific study summarized here, take a look at the original research article:
Ghosh, D. et al. Deep, noninvasive imaging and surgical guidance of submillimeter tumors using targeted M13-stabilized single-walled carbon nanotubes. Proc Natl Acad Sci 111(38), 13948-53 (2014).


Stephanie DeMarco (@sci_steph)
Staff Writer, Signal to Noise
PhD Candidate, Molecular Biology


References:

  1. “Ovarian Cancer: Signs and symptoms of ovarian cancer.” American Cancer Society. Last Modified February 4, 2016. http://www.cancer.org/cancer/ovariancancer/detailedguide/ovarian-cancer-signs-and-symptoms.
  2. Dolinsky C., Vachani C. & Bach C. “All About Ovarian Cancer.” Abramson Cancer Center of the University of Pennsylvania. Last Modified January 22, 2016. http://www.oncolink.org/types/article.cfm?c=203&id=8589.
  3. Buys, S.S. et al. Effect of Screening on Ovarian Cancer Mortality: The Prostate, Lung, Colorectal and Ovarian (PLCO) Cancer Screening Randomized Controlled Trial. JAMA 305(22), 2295-2303 (2011).
  4. Ghosh, D. et al. Deep, noninvasive imaging and surgical guidance of submillimeter tumors using targeted M13-stabilized single-walled carbon nanotubes. Proc Natl Acad Sci 111(38), 13948-53 (2014).
  5. Bashkatov, A.N. et al. Optical properties of human skin, subcutaneous and mucous tissues in the wavelength range from 400 to 2000 nm. Journal of physics. D, Applied physics 38(15), 2543 -2555 (2005).
  6. Thostenson, E.T., Ren Z. & Chou T. Advances in the science and technology of carbon nanotubes and their composites: a review. Composites Science and Technology 61(13), 1899-1912 (2001).
  7. Barone, P.W. et al. Near-infrared optical sensors based on single-walled carbon nanotubes. Nature Materials 4, 86 - 92 (2005).
  8. Chen, J. et al. SPARC Is a Key Regulator of Proliferation, Apoptosis and Invasion in Human Ovarian Cancer. PLoS ONE 7(8), e42413 (2012).