Tumour Cell Clusters V's individual Cells

Limiting potential for detachment of cell clusters which enter circulation.

Why adjusting current practice is urgent.

A very personal case in 2018/19 "my own mother" see Images

Page under development

Advances in circulating tumour cell (CTC) capture & numeration have confirmed that many cancer patients will have thousands of tumour cells in circulation at any point in time, yet patients never present with large numbers of secondary tumours.! For those paying attention to this specific field, the question was why?

Further more recent advances revealed that Tumour Cells which detach in clusters actually have the remarkable (and disturbing ) ability to unfold and pass through tiny vessels in a single-file chain by selectively retaining bonds at cell junctions. (Links to evidence & research below)

Saving lives and resources.

Seeking Progress to help encourage clinical practice to catch up with evidence demonstrating Tumour Cell Clusters being the main mechanism by which Cancers spread, as opposed to the common consensus that single detached tumour cells (CTC's) are. A very big deal due to the potential opportunities to help encourage adjustments to clinical practice & procedures to limit the potential for cluster detachment, and (as post surgery precaution) short period deployments of treatments which help break up any tumour cell clusters which may have entered vascular circulation (Lymphatic & Blood) after interventions. (Research Links below)

I believe if more Oncologists & related Clinicians took a little time to look at the research in this relatively new specific field , they could add their valuable experience to help make the case for forming a resourced team to urgently assess the potential of speeding progress by mining the large NHS data sets (patient case studies) on cancer patients. A team which could be expanded to bring together the small number of global experts in this specific field. while supporting efforts in R&D to limit potential for cluster detachment (Including by well meaning surgical interventions for tissue sampling), and assessment of treatments to breakup detached clusters into "individual cells" while in circulation.

Cancer Spread

The misleading common consensus!

The model above depicts single tumour cells entering circulation as a demonstration of how cancer spreads. Whats wrong with it? While Single cells do indeed frequently enter circulation "for reasons not yet clearly understood" they very rarely seed secondary tumours. If they did, tumours would be occurring in numbers akin to the spots that occur during Measles!

In recent years, the imperfect but cutting edge CTC Liquid Biopsy technology had confirmed that cancer patients will often have thousands of single tumour cells in circulation at any point in time, yet patients never present with thousands of tumours at once.!

Besides having spent years on CTC R&D with a very clever bio-med graduate. I'd also did a very simple but important bit of maths based on CTC data . Even with just 5 CTC's per 7.5ml of whole blood, an adult cancer patient (with 5L of blood) would have 3300 tumour cells in circulation at any point in time.

Note: depictions of single cell seeding can still be found on many of the main cancer websites.

Cancer Spread

Newer Important Observations on Clusters!

The original assumption was that Tumour cell clusters were too large to pass through narrow vessels to enter circulation & reach distant organs. Resent evidence from experiments like the above is just one example of how wrong that assumption was. With some tumours (the deadly ones), Tumour cell clusters have remarkably (and disturbing ) ability ability to unfold while selectively maintaining junctions to pass through capillary-sized constrictions in a single-file chain.

Like so much in cancer biology, the fascinating sub-set of processes enabling this require specific attention to detail. Knowing what this could mean for patient survival, trying to better understand those processes is one of the areas in my passionate interest in cancer biology which has kept me awake nights.

Remember: If single cell seeding had ever been a main mechanism of seeding, Secondary Tumours would have been presenting by the thousands (In their habitable sites) within Patients .

See additional research on Tumour Clusters below.

A very personal case in 2018/19 "my own mother"

Unfortunately at present the common consensus of individual CTC's having high metastatic potential remains wide spread in clinical oncology. That issue recently came to a head in a very personal case, in 2018 my own mother (the inspiration for the need driven quest "Since 2008" to overcome logistical limitations to progress on cancer) learned of a secondary local reoccurrence "a relatively large 3 cm tumour in the Left armpit Axillary Lymph Nodes" My mothers original tumour was in a breast which she had removed 10 years previous to this. Seemingly armed with that common consensus of individual CTC's having high metastatic potential, the treatment plan for my mother included "broad radiotherapy to the Chest" This was despite the fact that scans had returned no evidence of a potential tumour "seeding site" anywhere in the chest! When I asked questions of my mothers oncologist to try and understand the rationale, she wasn't pleased, and had mentioned single cells left behind at the original site (Chest Wall where the breast had been removed) as a potential seeding source for the secondary tumour.

Tried to point out that while I wasn't in any way professing to know all the answers, surely given a ten year opportunity for any cells left behind at the original site to have formed a tumour, & the recent removal of a 3cm tumour in the new secondary site (Armpit Axillary Nodes) that if the suspect seeding site for the main target area for radio therapy (Chest Wall where the breast had been removed) was correct, then surely medical imaging would likely have revealed a sizeable tumour growth in the chest wall too. Tried to explain that from our ten years of paying attention to the detail around CTC's & Clusters, that my mothers case highlighted something disturbing, but of potential value to progress in saving lives of other patients too.

Some numbered points on this.

1a/ Besides early tumour detection followed by wide margin removal of tumours, at present, the nature of the bonds between tumour forming cancer cells is likely among the biggest factors in whether a patient survives. and its by no means straight forward.

1b/ A big factor in the ability of Tumours to successfully Seed secondary Cancers is when cells break off/detach in clusters. As with tumours composed of cells maintaining tight junctions, those which usually only shed single cells into circulation are likely to be much less dangerous (In recent years CTC capture & enumeration provided evidence for this)

1c/ Urgent expansion in exploration of this could lead to immediate improvements in patient survival, reduce worry for some patients, (Knowing which Tumours are, less dangerous, or likely to reoccur) and if support could be gained for an ethically driven team to focus on these areas as part of Rapid Explore , it could eventually save great NHS financial resources too.

2a/ In cases like my own mothers here, If "on their own" the cell mutations leading to my mothers original tumour ever had high ability to readily detach clusters into circulation, (without mechanical disturbances during biopsies and surgery) given the time frame, she wouldn't have had survived. (Note; a single cubic mm of tumour tissue can easily contain more than 2million cells)

2b/ there is actually a high probability that her secondary tumours were seeded from cluster/s displaced during the unsuccessful attempt to remove all of the original primary tumour with wide margins (The Lumpectomy) or any one of the original biopsies (Multiple needle passes). Besides evidence of needle track seeding, evidence also exists of what had always been perfectly logical, mechanical interventions (even using fine needles) leave micro tears in tumour tissue! Support for urgently Mining NHS Data looking at similar cases could be useful for guiding treatment approaches for current and future patients. (The NHS partnership we wish to form could help in that too)

2c/ Given the the detail here, Potential induced spread, (Point 2a above) when my mother had a potential 3rd reoccurrence 2019 (head & tumour again occurring within the Lymphatic vascular network on the same side) and multiple points of evidence from various types of medical imaging, Ultrasound, PET, MRI. When monitoring and site specific targeting are not options, serious thought should be given to considering removal of the head and neck Tumour with wide margins (when possible). We first need a more a efficient NHS and more specialist skilled surgeons before being able to encourage more Lumpectomy's as first point interventions)

In short, my efforts to encourage reassessing the evidence before proceeding resulted in another unnecessary wasteful contest of 2018/19 which we could have done without. The common consensus of individual CTC's having high metastatic potential was a factor in that contest, as despite the mounting evidence that dethatched Clusters being the main seeding mode, and that single detached cells rarely seed secondary Tumours (The reasons for that should be a subject of great interest)

The mutated/damaged cells we call cancer.

Important note: not all cancers are equally as dangerous, big challenges remain our ability to make accurate assessments for many patients.

Cancer - Why no one has a complete picture.

Reasons include the limitations complexities & logistical challenges in each of the specific fields. Oncology is the branch of medicine that deals with the prevention, diagnosis, and treatment of cancers.

Cancer Pathology, Pathology Reports are currently the accepted standard for

Medical Imaging of tumours has came on along way, however some of the advanced scans remain expensive.

Pathologists are limited too due to technical limitations. Errors in in Histology have resulted in global scandals were cancers have been missed or over diagnosed. Yet it currently remains the main point of evidence (And requires invasive procedures to collect tissue samples)

There are lots of guides giving cancer patients advice on questions to ask their Oncologists. Samples of Guides on questions to ask: Macmillan, NCI, CRUK, Breast Cancer Care. However in practice, the very act of asking important question can go very wrong.

Some of the reasons as to why.

Firstly with current limitations, accurately gauging how dangerous one mutated cell line we call cancer is from another is often extremely limited. Making progress on those limitations was the motivation for devoting years to the background R&D, see Rapid Explore. While we don't claim to have anywhere near all the answers, after giving years to the background R&D we know we could be doing much better in several areas.

Despite the introduction of multidisciplinary teams in Cancer care, sometimes individual experts armed with their handbags of specific knowledge get upset when asked questions they don't know the answers too. Even worse when mistakes are made as opposed to informally conceding errors to proceed efficiently, the opposite can easily occur. Unfortunately by it's nature civil service management often lacks structure for progress, fear of scandal & tribal dynamics hinder the communication paths essential for progress.

Ideally a format for sensible informal communications (and when necessary informal apologies) would be in place, limiting unnecessary wasteful contests, encouraging more experts with knowledge specific to their fields to work together "professionally & respectfully" on assessing available evidence, helping drive progress & making the best rational assessments on treatment paths.

Below are some of the actual images in my own mothers case

Initial Static Images & PET Scan Report - A stressful time fraught with unnecessary contests hindering my ability to push for progress on getting support for Rapid Explore, costly delays to an ambitious endeavour which could eventually help improve the odds for many patients.

Ideally errors would be conceded without time wasted in unnecessary contests.

PET Scan Report

November 2018

The note on the PET report stating "Patient declined Adjuvant Chemotherapy" is another error, what was declined was "broad radiotherapy to the Chest"

Why? Besides the contraindications at the time, my mother had a mastectomy 10 years earlier, despite the eventual detection of a relatively large 3 cm tumour in the Left armpit Axillary Lymph Nodes, scans had returned no evidence of a potential tumour seeding site anywhere in the chest!

Basic Points

The logic

"Cell Clusters as the main mechanism" by which secondary cancers are seeded, as opposed to the common consensus of *Individual Cells " something that's actually a very big deal in saving lives!

1. It seems the first oncologist suspected the secondary 3cm tumour had been seeded from a Tumour source in the chest, however, If cells had indeed been left behind in the chest area after the mastectomy 10 years earlier, and with the known evidence point (the secondary local recurrence of a relatively large 3 cm tumour in the Left armpit) "if the seeding site of the secondary was in the chest" its reasonable to expect after that 10 year period that imaging would have detected a significant tumour in the chest That was not the case!

2. Although it remains a common consensus among many clinical oncologists & members of cancer teams that individual cells which detach from a primary Tumour are a main mechanism in how cancer spreads. We now know that individual detached tumour cells rarely result in seeding secondary cancers. That's also a massive deal when it comes to areas of focus in patient management, especially in relation to "limiting potential for cluster displacement" making use of treatments to break up detached clusters after surgical interventions & understanding metastatic pathways.

Why some cancers are much less dangerous than others, and how with some tumours which remain fairly benign (the bonds between the cells are such that the cells rarely detach in clusters) there is potential to inadvertently detach deadly clusters by well meaning interventions.(More detail available for cancer teams)

How I knew a bit about this. One of the challenging areas we'd been passionately working on for this past 10 years involved capturing live Circulating Tumour Cells (CTC's) , however around 6 years ago we began to suspect that although the efficient capture of live CTC's is a worthy goal in the quest for better Bio-markers & obtaining Patient Specific Mutations for pretesting treatments, (Reducing the guesswork) etc. For reasons not yet understood, single CTC's rarely result in seeding secondary Tumours. I also did a very simple but important bit of maths based on CTC data . turns out even with just 5 CTC's per 7.5ml of whole blood, an adult cancer patient (with 5L of blood) would have 3300 cells in circulation at any point in time.

If single cell seeding had ever been a main mechanism of seeding, secondary Tumours would have been presenting by the thousands (In their habitable sites) within Patients

Reducing the potential for Cluster Detachment & using drug treatments to break-up clusters which may detach "entering circulation" after some surgical procedures.

See below for some clear presentations from the small number of global experts in this field.

The remarkably ability of Tumour Cell Clusters to unfold while selectively maintaining junctions to pass through capillary-sized constrictions in a single-file chain. Due to the life saving potential in evolving better methods in managing patients, these are discoveries which should be given priority . At present i'm tied up with contests & trying to find a way to fund the launch of Rapid Explore, but if i survive and get there, I'll be putting a proposal to the NHS to put their large cancer patient data sets to use on this, and help form an urgent working group consisting of the small number of global experts in this field & any clinicians passionate about advances to assess the potential life saving implications.

Detail on GRHL2

https://www.genecards.org/cgi-bin/carddisp.pl?gene=GRHL2

Aggregates containing two or more CTCs (also known as circulating tumor microemboli; herein, CTC clusters) have been known for decades to seed colonies with greater efficiencies than individual CTCs (6⇓–8). Recent data suggest that clusters may have 50 times greater metastatic potential than individual CTCs (9). The greater colonization efficiency of CTC https://www.pnas.org/content/113/18/4947

Circulating tumour cell clusters pass through capillary-sized channels

Note: The wide section of the Microfluidic channel in the video (Left side) is about the with of a thick human hair.

Clusters of circulating tumour cells traverse capillary-sized vessels

Multicellular aggregates of circulating tumor cells (CTC clusters) are potent initiators of distant organ metastasis. However, it is currently assumed that CTC clusters are too large to pass through narrow vessels to reach these organs. Here, we present evidence that challenges this assumption through the use of microfluidic devices designed to mimic human capillary constrictions and CTC clusters obtained from patient and cancer cell origins.

Mohit Jolly (Rice U.): Circulating tumor cell cluster: A model for cancer metastasis

Excellent work by Dr. Mohit Jolly (Rice University Texas) on the potential value of quantifying GRHL2 to better gauge metastatic potential (Potential for cluster displacement) Imagine being able to tell some patients their tumour has low-seeding-potential "Something that current histology methods are often very poor at achieving, resulting in related missed diagnosis scandals, & over diagnosis. If combined with approaches William & i have been working on, this may reduce the potential for clusters being displaced into circulation & transform outcomes for some patients.

Extracts from source page http://www.cancermetastasislab.com/research/

We believe that targeting cell-cell junctions may represent a potent and previously under appreciated strategy to suppress CTC-clustering and cancer metastasis.

Dissecting fundamental mechanisms underlying cancer metastasis

Most recent findings

Using highly interdisciplinary approaches, we recently discovered key biological features of CTC clusters. For instance, we found that CTC clusters are characterized by hypomethylation of the binding sites for transcription factors that simultaneously regulate stemness and proliferation (such as OCT4, NANOG, SOX2 and SIN3A). Our results clearly link clustering and stemness features, and suggest that CTC cluster dissociation might be a valuable therapeutic strategy to reduce metastasis formation (Gkountela et al., Cell, 2019).

These results were featured on the cover of the journal “Cell” (see Figure 4).

Moreover, we discovered a new role for neutrophils in boosting the metastatic potential of CTCs. Particularly, we find that the association CTC-neutrophils enhances CTC proliferation through the release of specific cytokines. When this crosstalk is blocked, we observe a significant reduction in the metastatic spread of breast cancer cells (see Szczerba et al., Nature, 2019)

GRHL2 Gene protein encoder for transcription factor also see GRHL2 Human Protein Atlas

GRHL2 was already of interest in other studies - expanding on this is worthy of a team effort. if you have any input or thoughts that may be of help, please get in touch with Ian, details on the contact page.

Grhl2 reduces invasion and migration through inhibition of TGFβ-induced EMT in gastric cancer

GRHL2 in breast cancer