Growing tumors outside the body to kill the tumor still inside

To understand how to kill a tumor, you have to study the tumor. Historically, much of how scientists understand tumors comes from removing a tumor from a patient’s body, putting Cell_Cultureit in a plastic dish (called a petri dish), and studying whatever cells are grown in this dish. You may be familiar with the book “The Immortal Life of Henrietta Lacks” by Rebecca Skloot. This book talks about HeLa cells, which are cells that were taken from Henrietta’s cervical cancer, grown in a dish, and propagated for the past 60+ years as what is called a “cell line“.  These cells grow and divide indefinitely, and have been propagated and transferred from lab to lab to be studied.  HeLa cells are one of the most famous and most-researched cells that have helped scientists better understand cancer. HeLa cells are not the only cell line that exists or has been used to study cancer.  There are cell lines from lung cancer tumors, prostate cancer, brain cancer, and most other major cancers. However, there are a few problem with using cell lines to understand and treat cancer.

  1. Cell lines are EXTREMELY hard to create.  As you may imagine, a plastic dish is nothing like the environment inside the body that the tumor was removed from.  In the petri dish the cells are put into “media,”t he liquid that is used to feed the cells in the petri dish, and this media is also nothing like the nutrients and other growth factors feeding the tumor inside the body. Because of this unnatural environment, some of the tumor cells die – and in many cases mostor all of the tumor cells die.
  2. The cells that are left in the petri dish do not accurately represent the tumor anymore. A tumor isn’t a whole bunch of identical cells, but rather a tumor contains a lot of genetically different cells.  Scientists call this tumor heterogeneity. This is one of the reasons why drug resistant cells emerge after treating a tumor with drugs (like in the case of melanomadescribed in a previous post).  There are already drug resistant cells inside the tumor that don’t die when treated with drug.  Unfortunately, not all of these different cells in the tumor will live in a petri dish, so only a selected type or types of cells will live and can be studied.
  3. Even though cell lines had been the most useful tool in the past to understand cancer biology, they are not at all useful in understanding the EXACT tumor from a particular person. What does this mean? For example, drugs that kill HeLa cells in a petri dish might not work to kill another person’s cervical cancer because the genetic cause of that cervical cancer is different. In personalized medicine, the goal is to identify the drugs that will work to kill a particular patient’s tumor. Because of this, cell lines just aren’t good enough.

Scientists have been working on a number of solutions, and I’ll talk about four:

  1. Biobanking. A biobank collects excess tumor tissue from patients who are having a
    liquidnitrogenfreezers

    Where tumor tissue is stored in a biobank before researchers use it

    tumor removed as part of a surgery.  This tissue is immediately preserved by freezing and can then be used by researchers to study that particular tumor or many tumors of a particular type (e.g., lung cancer).  The disadvantage to this is that the tumor sample isn’t an unlimited resource. Once the tissue has been used up – it’s gone. The remaining examples all focus on growing the tumor tissue so that it can be propagated and used for many experiments.

  2. Modified cell line growth. HeLa cells were not grown in any special way, but researchers at Georgetown Universityhave found ways to grow tumor cells in a petri dish  that are identical to the tumor and nearly all tumors can grow under these conditions. So what are these conditions?  The researchers grow cells on top of a layer of mouse cells called feeder cells because they provide the cell-based nutrients to “feed” the tumor and allow it to grow.  They also use a particular inhibitor that allows the cells to grow indefinitely. They have created these modified cell lines from different types of tumors, from frozen biobanked tumors, and from as few as 4 live cells.  Even though this system, is better, it still doesn’t replicate the 3D architecture of a tumor…
  3. cancer organoids

    Cancer organoids. Notice the 3D clumps of cells after 217 days of growth. Thanks to the Kuo lab for the image

    Organoids. As you would expect the word to mean, an organoid is a mini 3D organ bud grown in a dish. Don’t imagine a teeny tiny beating heart.  These organoids are just clumps of cells, but an organized clump of cells that can help better understand cells and organs. The discovery of how to create organoids was so interesting that it was a 2013 Big Advance of the Year by The Scientists magazine. Scientist have also found a way to grow cancer cells into these 3D organoid structures. With tumor organoids, researchers can both study the genetics of the tumor (like you can with cell lines) as well as how the tumor behaved in a 3D environment that is more similar to what the tumor encounters in the body.  But what if we could do even better?

  4. Patient-derived xenograftsare when tumor tissue is taken directly from a patient’s tumor and put directly into a mouse.  Why would this be so awesome? The environment inside a mouse is more similar to the environment that the tumor is used to inside a person’s body.  The cells are less likely to die because they aren’t living in unnatural plastic. Also, a whole piece of tumor can be implanted into the mouse, maintaining the tumor cells connections to neighboring cells, which are critical for the tumor cells to communicate with one another for survival.

With all of these systems available to study tumors from a specific patient, what are scientists actually doing with these cells? In some cases, they are being used to sequence the genomes of the tumors to identify mutations that may be causing the tumor. If a tumor can be grown so that there is a lot of it, the tumor cells themselves can also be used to test treatments either in a dish or inside of a mouse. Imagine a cancer patient getting their tumor removed, part of the tumor is grown in one of the ways described above. Then the tumor is exposed to the top 10, or 50 or 100 anti-tumor drugs or combination of drugs to see what kills the tumor. This drug or combo of drugs can then be used to treat the patient. There are companies that are currently working on doing exactly this (check out Champions Oncology) so this “big dream” may soon become a cancer patient’s more promising reality.

 

What is that? A liquid nitrogen freezer

This is a new feature called “What is that?” where I will show a photo of something from the lab and then discuss what it is, what it does, and why scientists use it.  Every lab that I’ve ever worked in, I gave tons of tours – to my family, to other scientists, to kids and to the public!  Seeing what a real lab looks like and understanding how things work is one of the ways I use to demystify science and scientists. Think of this new series as a virtual lab tour with me as your tour guide!  I hope at some point soon to turn this feature into a short video series…but until then, enjoy learning about what “that” is!

So what are these? These are liquid nitrogen freezers.

liquidnitrogenfreezersLet’s start by talking about freezers in general. What temperature is your freezer at home? Well it needs to be cold enough to freeze things. The freezing temperature of water is 32° Fahrenheit (F) or 0° Celsius (C), so you want your freezer to at least be able to freeze water, but you also want it to preserve your food.  Part of preservation is preventing bacteria from growing since bacteria could potentially cause food poisoning when you thaw and eat the food (especially if it isn’t cooked first).  The colder the temperature, the less likely the bacteria is to be able to grow.  Also, the colder the freezer is, the less likely it is that enzymes (proteins that perform chemical reactions in cells) are active and breaking down the food’s nutrients. So the ideal freezing temperature for food is 0°F or -18°C.

In the lab, we have -20°C freezers that store reagents and look a lot like the upright freezer that my mom has in her basement to store extra food (and chill martini glasses). We also have -80°C freezers, also called ultra low freezers referring to the “ultra low” temperature.  These are better at storing proteins at a temperature that is cold enough to inactivate them. These ultra low freezers are also good for storing RNA.  RNA has a natural enemy, a protein called RNase, which is essentially a high-powered chomping Packman after the Pac-Dots dots that make up the RNA. At really cold temperatures, this RNase Packman is inactive and can’t work, keeping the RNA sample safe.

roses LN2

Creative Commons license. Original photo from Kasi Metcalfe on Flickr

As a bit of a segue, my 4 year old nephew calls me on Facetime a few times a month and always asks me to “show him some science.” One of the first things I showed him was our “Biospecimen Cryostorage” where we store the samples from our biobank, and which is what is pictured above. This room has three Liquid Nitrogen Freezers, and these are even colder that the ultra-low freezers. With liquid nitrogen (abbreviated to LN2) in the base, the whole tank is cooled to below -275°F or -170°C. If you feel like you’ve heard of liquid nitrogen before, you probably have. This is the liquid that is used at Science Centers to freeze roses and then smash them into a million pieces.  If you’ve seen any cooking show in the past 5 years, you’ve probably seen a chef use liquid nitrogen and pour it into a mix of cream and sugar to make a fast ice cream. However, in science we use liquid nitrogen as a way to quickly freeze and preserve living cells and tissue – a process overall known as cryopreservationLN2description

To better understand the photo and to really understand “what this is”, I have labeled areas above.  Each of the two tanks pictured can hold up to 40,000 individual samples.  These samples could be blood samples, tissue samples, samples of cells, etc.  In our case, these samples are stored in 2 ml (milliliter) tubes in plastics boxes in racks inside each freezer. Each box can hold 96 samples and each rack (see photo below) can hold 13 boxes (over 1200 samples). The freezers are automatically filled by tanks filled with liquid nitrogen sitting right next to each freezer.  Each freezer isn’t completely filled with liquid nitrogen for a few reasons. First, it would be really expensive but more importantly, it isn’t necessary. The temperature of liquid nitrogen is -190°C but the vapor keeps the tank colder than -170°C, which is still good enough to keep all of the proteins in these cells completely inactive.

LN2removalSamples are removed from these tanks on a regular basis and distributed to researchers.  You can see an image on the left of a rack of boxes being removed. It is important when selecting individual samples that they are not allowed to warm up and thaw.  This thawing can activate proteins that break down the samples or causes ice crystals when re-thawing that affect the integrity of a tissue sample or the viability of frozen cells, for example.

What is that? Liquid nitrogen freezers and a filler tank
What does it do? Keeps biospecimens or other samples at really low temperatures
Why do scientists use the? To keep samples well preserved and viable so that they work better for their experiments.

For more “What is That?”, click here.

What is a biobank? And why do scientists collect human tissue for research?

To understand what a biobank is and why they exist, it helps first to understand what type of “currency” is stored in the “bank” and what this “currency” will be used to purchase. Biobanks are a collection of “biospecimens” (the “currency”) and knowledge through research or preservation is often what can be “purchased” with this currency.  And as a quick FYI, biobank and biorepository are words that can be used interchangeably to describe the same thing.

Biospecimens can be any type of biological sample or material.  Seed biobanks, like the Svalbard Global Seed Bank in Norway, hold tens of thousands of seeds from over 4000 essential food crops.  The purpose of this biobank is to function as a back-up for seeds being stored in various countries. If a crop is wiped out because of disease or a zombie apocalypse, these seeds can be used to grow these crops again a biobank.

butterflies

photo credit Christian Guthier

The collection of birds, insects, butterflies, spiders and other animals and plants stored at the National Museum of Natural History at the Smithsonian Institute are also considered a biorepository.  There are a few reasons that these types of collections exist.  First, they are interesting to the public to show off the diversity of animals and plants found around the world and throughout time (I mean, who doesn’t like dinosaurs?).  But more than that, they are also immensely useful to scientists interesting in studying the animals themselves – including extinct animals, diversity, and evolution.

Biobanks that contain human tissue are most applicable to the study of humans and disease.  The biospecimens stored in these biobanks may include urine, blood, tissue (for example, extra tumor tissue removed during a surgery), feces (aka poop), cells (for example, cells scraped from the inside of the cheek or skin cells), cerebral spinal fluid, DNA or RNA.   The purpose of repositories that store tissue and fluids from people is to better understand diseases and use this understanding to develop molecular diagnostics and treatments. How is this done? Generally, scientists will compare biospecimens from many patients with a particular disease (for example, tumor tissue removed during surgery from patients with breast cancer or blood from patients with diabetes) to samples from patients who do not have that disease using one of those “-omic” analyses. Through understanding what causes the disease, methods can be devised to better detect the disease early or treatments can be developed to target the cause.

biobank_purpose

But you may be wondering how tissue biorepositories exist at all?  It is all because patients have been gracious enough to contribute some tissue, blood, skin, or nails for researchers to use in their research.  Biorepositories do not own this tissue and neither do the researchers – we are merely custodians of the tissue with the ultimate purpose to use the biospecimens for research.  The patient always comes first. Therefore, the first thing that is done before collecting tissue for research is to talk to the patient and explain why it is biobankers would like to collect and store their tissue.  The risks are explained and we ask for their permission through a process called informed consent (more on this in future posts).  If a patient does not consent to donate tissue for research, this does not affect their care in any way whatsoever.  It is the patients choice.  However, if they do agree to donate tissue, it will either be collected in the operating room (in the case of tumor tissue) or in pathology.  This tissue collection never disrupts medical care or diagnosis.  If all of the tissue is needed for diagnosing the patient, then that’s what happens and none is collected for research. Again, the patient and their medical care always come first.  If we are able to collect samples, they are stored in liquid nitrogen tanks until they are requested by a researcher.  We then make sure that the samples safely get to the researcher.biobank_workflow

Thousands (likely hundreds of thousands) of studies have relied on biospecimens to better understand the underlying disease in or to develop treatments.  For example, tissue from melanoma patients was used to identify a mutation found ~50% of patients who have melanoma.  This mutation can be specifically targeted by a drug that significantly improved progression-free survival in patients who typically have a dismal prognosis.  Even more studies on ongoing, with the goal of using knowledge gained from these priceless biospecimens to reach the promise of personalized medicine.