Weak GDP and Increasing Monetary Base

The GDP grew at a very weak rate. As seen above the Q1 2014 rate dropped to the lowest in several years. One can ascribe the sever winter, not global warming, for the drop, but still we have a significant concern.

M2 continues to rise so currency in circulation is strong but then again we have the FED backing by its continued QE efforts.

The Monetary Base also continues to rise. Banks are well off with reserves but none seems to leak into the economy to drive GDP

Comparing M2 and the Monetary Base we see the explosive growth of the MB while M2 maintains reasonable growth. Notwithstanding inflation is low.

Evil: Francis and Augustine

I have read Augustine and his Theory of Evil. It basically is the lack of good. But evil is a powerful word, a real powerful word. We call Hitler Evil. That has true meaning.

But today Francis tweets:

Inequality is the root of social evil.

With all due respect to The Bishop of Rome, does this mean all inequality is Evil? Some people run better than others, is that Evil? Some people make better investments than others. Is that Evil?

When one makes such a sweeping indictment then it often tends to reduce the strength of the word.

The Guardian jumped on this quickly. As they say:

But in last autumn's essay, Evangelii Gaudium, Francis wrote that: "Just as the commandment 'Thou shalt not kill' sets a clear limit in order to safeguard the value of human life, today we also have to say 'Thou shalt not' to an economy of exclusion and inequality. Such an economy kills … Today everything comes under the laws of competition and the survival of the fittest, where the powerful feed upon the powerless. As a consequence, masses of people find themselves excluded and marginalised: without work, without possibilities, without any means of escape. Human beings are themselves considered consumer goods to be used and then discarded." The claim that human beings have an intrinsic value in themselves, irrespective of their usefulness to other people, is one that unites Christianity and socialism. It can be even found somewhere in the shadows of Marxism, but there humans gain their value from history, and when they stand in its way, that's tough for them, as the millions of Stalin's victims could tell us. But if you think the market is the real world, it makes no sense at all, since in the market, value is simply the outcome of supply and demand.

Yes people have individual value, that is the essence of individualism.  Yet humans each have a duty to perform, to maximize their potential. Humans ultimately must answer for what they have made of their lives, either to themselves or God. Survival of the Fittest is a view of nature that has a basis in fact. It may not be a norm for social interaction.

I wonder what Augustine would think of this Bishop of Rome?

More Thoughts on Net Neutrality

There are two issues before the FCC this week. The Chairman's proposal for Net Neutrality, basically the "mail opening" proposal to charge the consumer for what is inside their mail, and the 3.5 GHz open band.

Consider if you will what may happen.

First, content providers are spread out across the world like polka dots on a map. They need to get to customers.

Second, there are many large broadband backbone entities that can get to the content providers but not to the customers. The largest are Level 3 and Google. Yes Google! Remember that fact.

Third, we poor customers need to get to that backbone so we subscribe at some extortionary rate to a local provider, say a cable company. The cable companies are now trying to lock us in and make us pay even more.

Fourth, the 3.5 GHz frequency band with 150 MHz of bandwidth is a shared band and could in effect bypass the cable guys and allow us to get in excess of 1 Gbps connect to that backbone out there. It could do so at a low cost. A very low cost. This is the basis of disintermediation, namely a new technology which changes the way things are done.

Fifth, thus the FCC may be sending a poison pill to the cable guys and there may eventually be the demise of these trolls.

Sixth, on the other hand if Google uses its backbone and if it captures this band then perhaps we have created a new AT&T pre-Divestiture.

I really wonder if anyone at the FCC has thought this through, or are even capable of doing this?

Le plus ca change!

Net Neutrality: The Simple View

The FCC was, is and always will be a highly political entity. It, in my opinion, will follow what the strongest winds of politics are at the moment. Having said this, one looks at the current Chairman's alleged proposal for Net Neutrality and in my opinion it appears to have been crafted by Comcast. I do not have any details, but in my experience with them, it smells that way.

Let us examine how the Internet works (see my paper on Net Neutrality).

1, There is a user and a provider. That is to say me and Netflix.

2. I pay my cable company to connect on either transmit or receive to and through them to an interconnection point in the Internet backbone. Not the cable provider's necessarily.

3. Netflix pays a similar backbone provider to connect from their servers to the backbone.

Thus there is a "long-distance" provider, say Level 3, and a local provider say Comcast.

Now Netflix pays Level 3 and I pay, in my case, Cablevision.

Now along comes Comcast and says they they will open my packets, my mail if you will, and if my mail comes from the wrong person they will delay it or charge me more. After all as the customer I pay no matter what.

Net Neutrality means one simple thing; all packets are equal and the cable guy, say Comcast, must not read my mail! After all they are not the NSA, yet they may be helping. But alas that is another story, I hope.

Thus Net Neutrality means that Netflix negotiates with Level 3 and I with my cable company. Period. And Comcast should not read my mail and interfere with my messages.

But don't count on that! Not with the folks we sent to DC this time!

ET Go Home

As NHPR states they have found ET in New Mexico desert. They state:

For decades, it was mere legend: an "Atari Dump" rumored to harbor millions of copies of E.T. The Extra-Terrestrial, a video game so bad that burying it in the New Mexico desert seemed the best way to move on. But now, the Atari graveyard has been exhumed, and the latest attempt to find the cache of game cartridges has been declared a success. Helped by heavy machinery, a crew found some of the games today, in a dig ...

I remember being at  Warner at the time, the Fall of 1982, and I was the Warner Cable interface with Atari. In October they were planning for a $2 billion 1983 and by December the roof collapsed. It was the best education in corporate planning gone amok. There was a flock of the most arrogant people I have ever met and then they were gone. Limousines gave way to the bus. Corporate jets gave way to People's Express.

I recall attending meetings and just wondering what was the basis for their original projections. The answer was always that is was always that way. As if change never occurs. It was one colossal mistake of arrogance after another.Perhaps a lesson for some, yet forgotten for many.

de Tocqueville and Piketty: A Contrast and Some Thoughts

Most people who are aware of de Tocqueville know of him solely by his writings on his journeys in the United States. There are two other writings worth reading, one on his journeys through Ireland and the forced slavery imposed upon the Catholics by the Crown, and just as important his understanding of the French Revolution. 

Unlike many others who wrote upon the French Revolution, one sees in de Tocqueville the same sense of “at handedness” (to use a Heidegger term) to pull together the collection of events and circumstances that led to the explosion which became the French Revolution. de Tocqueville is in sharp contrast to the intellectual depth of Israel, he does not bring forth the deep rooted philosophical elements emanating from the Enlightenment, but he details the day to day injustices done by the Government to the people at large. Wealth, the extreme wealth in France for some in pre-Revolution times is not even an issue. It fundamentally is the oppressive control of Paris, the seat of Government on all.

Piketty is almost the anti-de Tocqueville. Whereas de Tocqueville deals with the amalgam of small facts of daily life, the burdens placed upon the people under the presence of the Government, not just the King. de Tocqueville brings to his vision a proximity with the people, not the position of an academic hidden behind a wall of suppositions and beliefs.

Let me address a few key point in the order in which he presents them and they in a sense build the argument will shall make regarding Piketty. One should recall that de Tocqueville wrote his observations of America in 1835 and the reflections on the French Revolution in 1856, after the 1848 revolution again in France. Thus his observations of American reflect those of his youth and those here on the Revolution those just before he died.

Let us start with an observation of France before the Revolution in 1789.

“It was not until the beginning of the sixteenth century…that for the first time the right to work …came to be regarded as a “privilege” purchasable from the King.” (p 103)

The above details one of the major issues. Such rights, as that of choosing one’s profession, were no longer left to the individual but ascribed as a privilege purchased from the Crown. The Government could dictate, mandate, isolate, individuals based upon its determination of what in its mind and preference it saw and the desired outcome. The individual, no matter what they did, had little if any choice in what they did. The second key point is that it was purchasable. One could always find an entity of the Crown, the Government, to seek out and through a payment obtain what was sought. Thus one may ask if such is the case today in the United States? To a degree, yes. One can seek out the correct lobbyist, who has the correct Congress person, who can promote the desired legislation. The environs of Washington are now akin to a setting of camp followers, all seeking to provide “favors” for payment.

Individualism was not an unknown construct to de Tocqueville. He, to an extent, observed and coined the phrase as something most American. Yet as he explains below, individualism is not solely applicable to that lone person, it is seen even in France, as the ability and even desire of persons to seek satisfaction in the small, not just in the large.

“That word “individualism” which we have coined for our own requirements was unknown to our ancestors for the good reason that in their days every individual necessarily belonged to a group and no one could regard himself as an isolated unit. Nevertheless each of the thousands of small groups of which the French nation was then composed took thought for itself alone; in fact there was so to speak a group individualism which prepared men’s minds for the through-paced individualism which nowadays we are familiar.”  (p 96)

The most telling observation of de Tocqueville was the control over everything by Paris, by the Crown and its agents. All in France was measured, recorded, managed, and controlled via the Crow and its agents from Paris. The English had London but London did not do what Paris managed to do. Lyons was as controlled by Paris as was Bayeux, as was Annecy, as was Reims.

“Owing to this system of centralized information and controlling everything from Paris, a most elaborate machinery had to be set up for coping with the flood of documents that poured in from all sides and even so the delays of the administration were notorious…Not until the century’s end when the literary methods we associate with Rousseau and Diderot had begun to make good and affected the spoken language, did the rather maudlin sensibility affected by these authors creep into the style of our administration and even our businesses. Formerly so stiff and desiccated, our official style became unctuous…” (pp 62-63)

His second comment above is quite piercing, namely it is a backhanded swipe at Rousseau and Diderot, the Enclyclopedists. Unctuous indeed was the style, and maudlin was the sense.

Then as stated below is the style of the revolution. He does not take to compare it to America, but to examine it as an act of academic investigation, leaving the details as exercises for the student.

“When we closely study the French Revolution we find that it was conducted in precisely the same spirit as that which gave rise to so many books expounding theories of government in the abstract. Our revolutionaries had the same fondness for broad generalizations, cut-and-dried legislative systems, and a pedantic symmetry; the same contempt for hard facts; the same taste for reshaping institutions on novel, ingenious, original lines; the same desire to reconstruct the entire constitution according to the rules of logic and preconceived system instead of trying to rectify its faulty parts.” (p 147)

The telling point above is the contempt for hard facts. The French academic all too often enjoys the mental gymnastics, devoid of the facts that the Scot would bring. The French Enlightenment and the Scottish Enlightenment were revolutions in thought that were at the same time contradictions. Ideas devoid of facts versus Facts embodying ideas. The contrast is Rousseau versus Smith. The issue of rectifying faults is the key refrain. The current best example is the ACA in the United States. Instead of rectifying the faults, most of which were well known, the body politic reshaped reality. The result may very well be the same as that of the intellects in the Revolution.

“I have sometimes asked Americans whom I chanced to meet in their own country or in Europe whether in their opinion religion contributes to the stability of the State and the maintenance of law and order. They always answered, without a moment’s hesitation, that a civilized community, especially one that enjoys the benefits of freedom, cannot exist without religion. In fact, an American sees in religion the surest guarantee of the stability of the State and the safety of individuals. This much is evident even to those least versed in political science.” p (153)

Now de Tocqueville takes on the Economists, a new type of character whose entry in the eighteenth century foreshadowed this trade’s power over many of our current debates.

“Towards the middle of the eighteenth century a group of writer known as the …”Economists” who made the problems of public administration their special study, came to the scene. Thought the Economists figure less prominently than our philosophers in histories of the period and perhaps did less then they towards bringing about the Revolution, I am inclined to think it is from their writings that we learn most of its true character…Their chief targets of attack were those institutions which the Revolution was destined to sweep away forever…” (p 158)

Finally we examine his examination of the ruling passions of the French. He states:

“Readers of this book who have followed carefully my description of the eighteenth century France will have noticed the steady growth amongst the people of two ruling passions…One of these, the more deeply rooted and long-standing was the intense, indomitable hatred of inequality….The other ruling passion, more recent and less deeply rooted, was a desire to live not only on an equal footing but also as free men.” (pp 207-208)

The two passions were the drivers of the people; the hatred of inequality and the desire to be free. France looks at Liberty, Equality and Fraternity. Yet when one examines Equality one sees not equality of opportunity but the inequality of a class based society as the concern, as the desire of the people. The inequality was that certain people had privileges perforce of who they or their ancestors were. They were not equal before the law; there was rampant inequality, many getting to the head of the line while many were left behind.

Equality in America is the equality of individualism, that all people are equal before the law, that there is no privilege, that each individual can do with their talents as much as any other and that success is predicated on how well those talents are valued in the market. To the American in de Tocqueville’s period, the Government in America empowered the individual, while to the Frenchman at the time of the French Revolution the Government established mass areas of inequality. Americans cherished equality whereas the French detested inequality. This I believe is a core element in the economic politics of Piketty.

New Bandwidth at 3.5 GHz

The FCC has announced its intent to release some 150MHz of bandwidth at 3.5 GHz. They state:

The Federal Communications Commission today took steps to provide more spectrum for general consumer use, carrier-grade small cell deployments, fixed wireless broadband services, and other innovative uses, through the creation of a new Citizens Broadband Radio Service. The Commission  proposed rules for the Citizens Broadband Radio Service in a Further Notice of Proposed Rulemaking that advances the Commission’s efforts to meet the growing demand for spectrum by  proposing to make 150 megahertz available in the 3.5 GHz Band. The FNPRM proposes innovative spectrum sharing techniques to unlock the value of the spectrum between 3550 MHz and 3650 MHz, and seeks comment on extending the proposed service to 3700 MHz. Specifically, the FNPRM proposes a three-tiered access and sharing model comprised of federal and non-federal incumbents, priority access licensees, and general authorized access users. Together, the proposals seek to promote flexibility and innovation by leveraging advancements in technology to facilitate sharing between different users and uses, including incumbent government uses.

This may represent a significant opportunity for drastic expansions of broadband. Using MIMO and OFDM alone means well in excess of 1 Gbps capabilities. The only weakness is that propagation at 3.5GHz is line of sight.

This proposal is a result of a PCAST Report. The Report proposes (also see dailywireless piece on this):

Today’s spectrum users are in three categories: Federal users, licensed commercial users, and unlicensed commercial users. The proposed system will add three new categories. The first is Federal Primary Access, for legacy Federal users that share their spectrum on a first priority basis with other Federal users or commercial users. Conflict is managed by registering spectrum usage in a database. Secondary Access users are Federal or commercial users that have the next priority to shared Federal spectrum. Applications that require higher power and better quality of service than today’s unlicensed devices will benefit from this category, although a fee may be required to access this spectrum. The third category, General Authorized Access, has the lowest priority, and supports less critical low power applications such as meter reading or entertainment.

We have proposed a more robust real time approach almost a decade ago when examining Software Defined Radios and had held discussions with FCC members at the time.

In fact our proposal allowed for real time adaptive spectrum management in a real time bidding approach. The bidding approach also mimicked a Bitcoin payment system. It is good to see that some of these ideas have some legs, albeit very slowly. Then again it is the FCC after all.

Obesity: Disease, Genes, or Choice?

There is an ongoing battle over obesity. The AMA decided it was a disease, something that "happens", perhaps due to what a human does, but it is not the sole result of a human decision. It is like mumps, when you go to school and the other kids have it, you get exposed, and blow up. Some look for the genetic link. The refrain, "I can't help it, it is in my genes." has been sounded again and again. Then to my surprise in Nature there is a call for personal responsibility, namely they just eat too much.

The author states:

Obesity is an important contributor to the prevailing burden of chronic disease, lying on the causal pathway to much of what plagues modern society and its people — heart disease and diabetes to name two of the most serious. However, not only can these diseases develop in the absence of obesity, but not everyone with a high BMI develops any such condition.The categorization of obesity as a disease could have a pernicious influence on efforts to remedy the problem at its true origins. The treatment of diseases customarily involves drugs, medical technology, clinic visits and surgical procedures. If obesity is a disease, the therapeutic advances on which its management depends presumably reside in these domains.

 Again, for almost all people, for every 3500 excess kcal we gain a pound. If we burn 2000 kcal per day and we eat 2500 kcal per day, that is three 12 oz sodas, then we can gain a pound a week, or 52 pounds a year!

Thus it is easy to become obese. Frankly it is just as easy to reduce that process, possibly a little slower, but it can be revered.

Thus it is good to hear a voice which lays the problem at the proper doorstep.

Google, Fiber and the Franchise

There has been some recent talk of Google and its fiber Odyssey. In a recent ARS Technica piece they discuss the possibility. Having done some New York builds in my time and being still somewhat aware of the process, at no time does anyone seem to address the issue of the Franchise. What do they expect. Just start digging holes, pull the fiber and well? In New York. Ever head of IBEW Local 3? If not then you better learn quickly. You just don't send a team from Palo Alto into the city and pull whatever and wherever.

As the article mentioned above states:

Google recently announced that it chose nine metro areas around the country for potential Fiber deployments. The closest ones to New York City are Raleigh-Durham in North Carolina and Atlanta, Georgia. New York City already has fiber in the form of Verizon FiOS, and Google has focused mostly on underserved areas where municipal officials are willing to provide expedited permitting and other perks. There are still millions of Americans without broadband, so there are plenty of areas where Google Fiber is needed. One thing that is clear is that Google is building up its Fiber team. Job listings indicate that more than 60 positions are open. There is one other Google Fiber position open in New York, for a network infrastructure design manager.

 But assuming you break bread with the Union types, a real big assumption, then what of the Franchise? That may take nearly forever. You can bet that if Comcast gets Time Warner that any chance another entity has of doing anything is zero, I have been there.

Lastly, the process of getting a Franchise may very well take forever. The costs are unbelievable. How then can one get any return on investment? That is the key question.

Also, if wireless keeps doing what it is doing and expanding data rates and lowering costs, then why build fiber at all?

The International Business Times lays out a more complete tale. They state:

Underneath Manhattan lies a vast labyrinth of tunnels that was originally built for telephone wire after the Great Blizzard of 1888. It runs from all the way from downtown Manhattan to the Bronx, and it's controlled by Empire City Subway (ECS), a Verizon subsidiary. Verizon claims that it maintains the tunnels, and it points to its own fiber-optic FiOS network as proof. But critics, including one of Verizon’s competitors, as well as other businesses that lease the space to run their own cables through there, recently told Crain’s New York that the tunnels tell a different story:

"Conduits are filled with cables from defunct Internet providers that went belly-up after the dot-com bust in 2000. Verizon itself left severed copper wire in lower Manhattan ducts after installing a fiber-optic network following Superstorm Sandy. (The company says the cables could be easily removed, if needed.)"

The conduit system that could supply New York with Google Fiber is a crowded mess, which is unlikely to change in the short term. Why would Verizon clear the way for its competition?

 Indeed, there are a plethora of obstacles. First the Franchise. We wrote of our recent experiences. That process is endless, meeting after meeting with every citizen having a say. Second is rights of way as discussed above. The incumbent has those rights, not the city. Try and displace them. Third is as mentioned above is the unions. New York makes Afghanistan look like the paradigm of correctness. I suspect there are unions to manage the "Porta Johns". Fourth, is the process of getting permits for this and that. Those who succeed in Real Estate have spent decades mastering this effort. A new guy on the street just cannot master the effort.

But remember the key factor. Wireless now is a winner. OFDM allows 10 bps/Hz, add to it adaptive beamformed antennae and we may get another factor of 5 to 10. Then HDTV can be compressed to 4 Mbps. Thus we can achieve a Gbps speed per user and can send a ton of video, which Verizon already has access to via FIOS. Ever wonder why they abandoned FIOS?

Getting a Job

I am always amazed when I read something about Google, namely is it true or just for the show. I recall being out there visiting the Chairman in the old building which I believe was formerly an Atari facility in one of my prior lives.

But as is stated in today's NY Times by one of the commentators who frequently tries to opine on the technology space, all too often in my opinion with little understanding, Google tries to hire people with certain skills. He states:

...the first thing Google looks for “is general cognitive ability — the ability to learn things and solve problems,” he said. In that vein, “a knowledge set that will be invaluable is the ability to understand and apply information — so, basic computer science skills. I’m not saying you have to be some terrific coder, but to just understand how [these] things work you have to be able to think in a formal and logical and structured way.” But that kind of thinking doesn’t have to come from a computer science degree. “I took statistics at business school, and it was transformative for my career. 

Analytical training gives you a skill set that differentiates you from most people in the labor market.”

A lot of work, he added, is no longer tied to location. “So if you want your job tied to where you are, you need to be: A) quite good at it; and B) you need to be very adaptable so that you have a baseline skill set that allows you to be a call center operator today and tomorrow be able to interpret MRI scans. To have built the skill set that allows you to do both things requires a baseline capability that’s analytical.”

The overall discussion is how to get a job at Google. Perhaps it should have been how to get a job period. Now just what the second paragraph above intends to say is too complex for me. Just what does he mean being tied to a location. Back in the 60s we moved every other year, from Boston to New Jersey to Boston, to DC, to Chicago, to Atlanta. Frankly I wonder if this is what he is saying. Then the ability to be a call center operator and a Radiologist is a non sequitur of the highest level. I guess it is just what one would expect from HR and a reporter.

Having just returned from a week trip with grandson number 2 to five colleges in anticipation of his next step, the key issue is what is he doing to get a job? He may still be a High School Freshman but now is the time to start that discussion. He may want at this stage to be a Civil Engineer, a noble calling, but then at his age I wanted to be a jet pilot, not knowing that at 6'3" I most likely would lose my head if ever ejected. But the earlier one starts the better is the process. He will not get a job as an anthropologist, there are very few of them, unless you are self funded by a large trust fund. Yet there is a continual demand for Civil Engineers, and Chem Es as well. 

Thus prior planning does indeed prevent poor performance. It is not just analytical thinking but doing so in a long term perspective, looking forward to have skills which are portable, marketable, and sellable. An electrician always has a better chance that an anthropologist.

καλα πασκα

Happy Easter!

Methylation and Ancestors

Every time we learn more about genes and their operations we add complexity. Epigenetics has added a dimension which oftentimes surpasses much of what we have learned before. In a recent Science article the authors examine the epigenetic differences in humans and their ancestors. They state:

Ancient DNA sequencing has recently provided high-coverage archaic human genomes. However, the evolution of epigenetic regulation along the human lineage remains largely unexplored. We reconstructed the full DNA methylation maps of the Neandertal and the Denisovan by harnessing the natural degradation processes of methylated and unmethylated cytosines. Comparing these ancient methylation maps to those of present-day humans, we identified ~2000 differentially methylated regions (DMRs). Particularly, we found substantial methylation changes in the HOXD cluster that may explain anatomical differences between archaic and present-day humans. Additionally, we found that DMRs are significantly more likely to be associated with diseases. This study provides insight into the epigenetic landscape of our closest evolutionary relatives and opens a window to explore the epigenomes of extinct species. 

This is an interesting first step well worth the following!

One suspects that the more we understand methylation, miRNAs etc the better we can understand some of the vagaries of life.

QALYs and "Death Panels"

One of the major controversies on the ACA roll out was the use of QALYs as a means to ascertain what treatment was best. A QALY is a British construct to control NHS expenditures. Thus if you are say 75, and have prostate cancer, then the NHS may very well deny you any treatment. After all you are dead in 7 years anyway, and so you may die a bit quicker it is cheaper than way. That in a way is a "Death Panel" After all it does not take into account you as an individual and your individual state of health. You are deemed to be part of a group of 75+ year old males.

Now the ACA was to have prohibited that. Yeah, that is what they said. But today the NY Times has a piece that says that although the ACA does not expressly state that the "Societies" are doing it on their own.

They state:

..., a visiting scientist in the ethics department at the National Institutes of Health, said the move by some societies to incorporate economic analysis “heralds an important shift in the way doctors in America are talking about cost and value.”

He said that having societies do such evaluations was better than having a doctor make such trade-offs while treating an individual patient, which is sometimes called bedside rationing.

Still, it is unclear if medical societies are the best ones to make cost assessments. Doctors can have financial conflicts of interest and lack economic expertise.

The cardiology societies, for instance, plan for now to rely on published literature, not commission their own cost-effectiveness studies, said ... a professor at Stanford and co-chairman of the committee that wrote the new policy.

They plan to rate the value of treatments based on the cost per quality-adjusted life-year, or QALY — a method used in Britain and by many health economists.

The societies say that treatments costing less than about $50,000 a QALY would be rated as high value, while those costing more than $150,000 a QALY would be low value. “We couldn’t go on just ignoring costs,” ... said.

So the Dems in DC are not expressly killing off the older folks they have apparently gotten certain allies in Health Care to do the dirty task for them.  I have written extensively on QALY analysis and it is filled with many bias elements, especially against health older men. So do we read a message here or what? 

CRISPR Patent

The PTO had issued a patent on CRISPR to Broad and MIT. This is the beginning of something interesting.

Filed 15 October 2013 and Issued 15 April 2014! The PTO has not moved this fast since Edison!

The prime claim is:

What is claimed is:

1. A method of altering expression of at least one gene product comprising introducing into a  eukaryotic cell containing and expressing a DNA molecule having a target sequence and encoding the gene product an engineered, non- 5 naturally occurring Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)—CRISPR associated (Cas) (CRISPR-Cas) system comprising one or more vectors comprising:

a) a first regulatory element operable in a eukaryotic cell 10 operably linked to at least one nucleotide sequence encoding a CRISPR-Cas system guide RNA that hybridizes with the target sequence, and

b) a second regulatory element operable in a eukaryotic cell operably linked to a nucleotide sequence encoding a 15 Type-II Cas9 protein,

wherein components (a) and (b) are located on same or different vectors of the system, whereby the guide RNA targets the target sequence and the Cas9 protein cleaves the DNA molecule, whereby expression of the at least 20 one gene product is altered; and, wherein the Cas9 protein and the guide RNA do not naturally occur together.

Prostate Cancer: Genetics and Epigenetics

There has been an explosion in genetic “causes” for many cancers and prostate cancer, PCa, is not the exception. We have completed a White Paper which covers the material herein in some detal.

One of the most significant factors has been the ability by some to take metrics of multiple gene expressions and allege that with the proper weightings these single dimensional metrics are prognostic. The problem with the metrics is often that they do not relate to actual genetic control mechanisms. We consider here an example in PCa of several genes and miRNAs which taken together create a putative malignant state.

Specifically we examine three elements:

1. p53, the classic oncogene which is a control element for keeping cells in a homeostatic state and avoiding malignant changes.

2. miRNA 34, or miR-34 which is a micro RNA and is also found to have a controlling effect upon a cell.

3. MET, a tyrosine kinase receptor which can be activated by HGF, the hepatocellular growth factor ligand, and which can activate multiple pathways and if activated and done so in an uncontrolled manner can result in malignancies.

This examination is predicated on a recent paper by Cheng et al (2014) which discusses the joint regulation effects of p53 and miR-34.

This section discusses the micro RNA process and its impact on PCa. Micro RNAs, miRNA, are small single stranded RNAs which when in the cytoplasm may often bind to other RNA on complement binding sites and thus change or incapacitate the mRNA to which it binds from being translated into a protein. Craig Mello was awarded the Nobel Prize in 2006 for the discovery and his Nobel Lecture provides an excellent overview of the early stages of miRNA investigation.

In a recent paper by Cheng et al (2014) they state:

The miR-34 family was originally found to be a direct target of p53 and is a group of putative tumor suppressors. Surprisingly, mice lacking all mir-34 genes show no increase in cancer formation by 18 months of age, hence placing the physiological relevance of previous studies in doubt.

Here, we report that mice with prostate epithelium-specific inactivation of mir-34 and p53 show expansion of the prostate stem cell compartment and develop early invasive adenocarcinomas and high-grade prostatic intraepithelial neoplasia, whereas no such lesions are observed after inactivation of either the mir-34 or p53 genes alone by 15 months of age.

Consistently, combined deficiency of p53 and miR-34 leads to acceleration of MET-dependent growth, self-renewal, and motility of prostate stem/ progenitor cells.

Our study provides direct genetic evidence that mir-34 genes are bona fide tumor suppressors and identifies joint control of MET expression by p53 and miR-34 as a key component of prostate stem cell compartment regulation, aberrations in which may lead to cancer

This is a murine model which putatively demonstrates that a blocking of both miR-34 and p53 leads to PCa. Specifically, this is MET pathway dependent growth.

As noted in Bioscience Technology[1]:

Previous research at Cornell and elsewhere has shown that another gene, called p53, acts to positively regulate miR-34. Mutations of p53 have been implicated in half of all cancers. Interestingly, miR-34 is also frequently silenced by mechanisms other than p53 in many cancers, including those with p53 mutations.

The researchers showed in mice how interplay between genes p53 and miR-34 jointly inhibits another cancer-causing gene called MET. In absence of p53 and miR-34, MET overexpresses a receptor protein and promotes unregulated cell growth and metastasis.

This is the first time this mechanism has been proven in a mouse model, said Alexander Nikitin, a professor of pathology in Cornell’s Department of Biomedical Sciences and the paper’s senior author. Chieh-Yang Cheng, a graduate student in Nikitin’s lab, is the paper’s first author.

In a 2011 Proceedings of the National Academy of Sciences paper, Nikitin and colleagues showed that p53 and miR-34 jointly regulate MET in cell culture but it remained unknown if the same mechanism works in a mouse model of cancer  (a special strain of mice used to study human disease).

The findings suggest that drug therapies that target and suppress MET could be especially successful in cancers where both p53 and miR-34 are deficient.

Also, the number of stem cells in mice with both p53 and miR-34 silenced increased substantially compared with control mice or mice with only miR-34 or p53 independently silenced.

“These results indicated that together [miR-34 and p53] regulate the prostate stem cell compartments,” said Nikitin.

This is significant, as cancer frequently develops when stem cells become unregulated and grow uncontrollably, he said.

Researchers further found that p53 and miR-34 affect stem cell growth by regulating MET expression. In absence of p53 and miR-34, MET is overexpressed, which leads to uncontrolled growth of prostate stem cells and high levels of cancer in these mice.

From Tang’s Lab at MD Anderson we have[2] (see Liu et al):

Cancer stem cells (CSCs), or tumor-initiating cells, are involved in tumor progression and metastasis1. MicroRNAs (miRNAs) regulate both normal stem cells and CSCs and dysregulation of miRNAs has been implicated in tumorigenesis6. CSCs in many tumors—including cancers of the breast, pancreas, head and neck, colon, small intestine, liver, stomach, bladder and ovary—have been identified using the adhesion molecule CD44, either individually or in combination with other marker(s).

Prostate CSCs with enhanced clonogenic17 and tumor-initiating and metastatic capacities are enriched in the CD44+ cell population, but whether miRNAs regulate CD44+ prostate cancer cells and prostate cancer metastasis remains unclear. Here we show, through expression analysis, that miR-34a, a p53 target was underexpressed in CD44+ prostate cancer cells purified from xenograft and primary tumors.

Enforced expression of miR-34a in bulk or purified CD44+ prostate cancer cells inhibited clonogenic expansion, tumor regeneration, and metastasis. In contrast, expression of miR-34a antagomirs in CD44− prostate cancer cells promoted tumor development and metastasis. Systemically delivered miR-34a inhibited prostate cancer metastasis and extended survival of tumor-bearing mice.

We identified and validated CD44 as a direct and functional target of miR-34a and found that CD44 knockdown phenocopied miR-34a overexpression in inhibiting prostate cancer regeneration and metastasis. Our study shows that miR-34a is a key negative regulator of CD44+ prostate cancer cells and establishes a strong rationale for developing miR-34a as a novel therapeutic agent against prostate CSCs.

Overall we examine here a four part set of elements related to PCa; receptors, pathway elements, mi RNAs and methylation. We outline this graphically below:

Note that in the above each plays a role in the development of PCa.

This has been known for a while. We see in Yamamura et al (2012) that they observed:

MicroRNA-34a (miR-34a), a potent mediator of tumor suppressor p53, has been reported to function as a tumor suppressor and miR-34a was found to be downregulated in prostate cancer tissues. We studied the functional effects of miR-34a on c- Myc transcriptional complexes in PC-3 prostate cancer cells. Transfection of miR-34a into PC-3 cells strongly inhibited in vitro cell proliferation, cell invasion and promoted apoptosis. Transfection of miR-34a into PC-3 cells also significantly inhibited in vivo xenograft tumor growth in nude mice. miR-34a downregulated expression of c-Myc oncogene by targeting its 39 UTR as shown by luciferase reporter assays. miR-34a was found to repress RhoA, a regulator of cell migration and invasion, by suppressing c-Myc–Skp2–Miz1 transcriptional complex that activates RhoA.

Overexpression of c-Myc reversed miR-34a suppression of RhoA expression, suggesting that miR-34a inhibits invasion by suppressing RhoA through c-Myc. miR-34a was also found to repress c-Myc-pTEFB transcription elongation complex, indicating one of the mechanisms by which miR- 34a has profound effects on cellular function. This is the first report to document that miR-34a suppresses assembly and function of the c-Myc–Skp2–Miz1 complex that activates RhoA and the c-Myc-pTEFB complex that elongates transcription of various genes, suggesting a novel role of miR-34a in the regulation of transcription by c-Myc complex.

It is interesting to see that we have a miRNA as a tumor suppressor. It is a key change in the way we can understand the overall pathway control paradigm. Thus the miRNA acts in a powerful manner to modulate cell growth and proliferation.

Micro RNA

The development of our understanding of micro RNAs has evolved from that of elements just left over to key control factors in major pathway expression. From Pekarik et al:

Among all previously described factors involved in the initiation and development of prostate cancer another element interconnecting several cellular processes may be traced. This element is represented by microRNAs (miRNAs), short non-coding regulatory molecules involved in multitude of processes in eukaryotic cells. They play a role in virtually each step of tumour formation and progression. miRNAs networks affect apoptotic pathways, cellular growth, responsiveness to growth factors and anticancer drugs, inhibit expression of tumour suppressor genes or permit expression of oncogenes.

Classical textbooks refer to carcinogenesis as a harmonic process caused by a loss of function of tumour suppressor genes and simultaneous activation of oncogenic genes. Recent progress in miRNAs function studying did not change this definition substantially; it only extended our understanding of regulation of this intrinsic network by miRNAs which can be likewise characterised as oncogenic miRNAs and antitumour miRNAs.

Indeed, we now see that tumor growth is a highly complex amalgam of genetic elements and supra-genetic elements as well. We have also argued that in many cases we see extracellular matrix interactions as well as free radical excitation of cells as well.

Oncogenic miRNAs are those that directly or indirectly suppress gene expression of tumour suppressors or proapoptotic genes and vice versa anti-tumorigenic miRNAs are those that reduce expression of oncogenic proteins. miRNAs are involved in nearly all types of cancer studied so far and they target classical oncological pathways. However, certain miRNAs were specifically associated with defined tumour types suggesting that they are involved in specific processes related to a cancer type or a tissue of origin. With regard to the number of genes regulated by miRNAs it is not surprising that these small regulatory molecules play a role also in the resistance of cancer cells to various anti-cancer drugs. In that respect, miRNAs become very attractive target for potential therapeutic interventions.

Recent research has revealed existence of miRNAs circulating in human blood serum. More surprisingly, it was found that levels of various miRNAs are altered in response to various physiological changes and some of these changes are well correlated with tumour existence. This makes circulating miRNAs a very attractive non-invasive cancer biomarker.

miRNAs have come to the fore as one of the several epigenetic factors which can precipitate various malignancies. The added factor of methylation as a silencing mechanism also adds to but further complicates the understanding of cancer progression. Thus, when we see loss of a miRNA, we may actually be indirectly observing the effects of methylation of the CpG region about that miRNA encoding region.

The relationship between miRNAs and pathway control elements is now being better understood. From Yamamura et al:

MicroRNAs (miRNAs) are highly conserved, single stranded, non-coding RNAs of approximately 22 nucleotides that regulate gene expression by posttranscriptional silencing of specific target mRNAs, by repressing translation or cleaving RNA transcripts. miRNAs regulate diverse cellular processes such as cell-cycle progression, proliferation, apoptosis and development. miRNAs have been shown to function as oncogenes or tumor suppressor genes.

The p53 tumor suppressor is deleted or mutated in more than 50% of human tumors and is a key molecule which suppresses malignancies. p53 has been found to target the miR-34 family and the ectopic expression of miR-34 genes has drastic effects on cell proliferation and survival. Ectopic miR-34a causes cell-cycle arrest in the G1 phase and apoptosis. As p53 has been found to target miR-34a and since, cell-cycle arrest and apoptosis are also end points of p53 activation, the miR-34a gene may be a mediator of p53 function. The proto-oncogene c-Myc regulates cell proliferation and transformation both transcriptionally and non-transcriptionally and is frequently deregulated in human cancers

miR-34 is one of now hundreds of micro RNAs, which are short, generally 22 base pairs, and non-coding RNA segments. They are now well known as control elements in the expression of genes and have significant control mechanisms.

miR-34 Structure

From NCBI[3] (1p36.22; 1p36.22):

microRNAs (miRNAs) are short (20-24 nt) non-coding RNAs that are involved in post-transcriptional regulation of gene expression in multicellular organisms by affecting both the stability and translation of mRNAs. miRNAs are transcribed by RNA polymerase II as part of capped and polyadenylated primary transcripts (pri-miRNAs) that can be either protein-coding or non-coding.

The primary transcript is cleaved by the Drosha ribonuclease III enzyme to produce an approximately 70-nt stem-loop precursor miRNA (pre-miRNA), which is further cleaved by the cytoplasmic Dicer ribonuclease to generate the mature miRNA and antisense miRNA star (miRNA*) products. The mature miRNA is incorporated into a RNA-induced silencing complex (RISC), which recognizes target mRNAs through imperfect base pairing with the miRNA and most commonly results in translational inhibition or destabilization of the target mRNA.

There has been a great amount of research regarding the impact of miRNA on cancer and especially on PCa. miRNAs may downregulate tumor suppressor genes such as PTEN. This has been seen in miRNA 21. Colin and Croce have provided several review article regarding miRNA and their influence on cancers. They argue that miRNA alterations are heavily involved in the initiation of many cancers. Their focus had been on CLL, chronic lymphocytic leukemia, and its initiating miRNAs, miR 15 and miR 16. Coppola et al (2010) provide a detailed summary of miRNAs and PCa.

For example miR34 can cause the activation and recapitulate p53 which in turn induces cell cycle arrest and apoptosis. Loss of the miR34 can result in the impairment of the p53 control of apoptosis and permit the cells to proliferate. Coppola et al perform a detailed analysis of all of the above related miRNAs and their resultant impact on PCa. miR-21 up-regulation leads to PTEN loss and thus is an oncogene.

Recent work by Poliseno et al has shown that PTEN can be down regulated via miR-106b. It had already been known that PTEN could be down-regulated by miR-22, miR-25 and miR-302. Their work demonstrated that miR-22 and miR-106b are overexpressed  in PCa miR-106b is an intronic miRNA. The work of Poliseno thus has demonstrated a proto-oncogenic miRNA dependent network that regulates PTEN and thus can have a significant role in initiating PCa.

Micro RNAs are regulators of mRNA, the post transcriptional result which is then used to generate via translation the operative protein. Currently there are nearly 1,000 identified miRNAs. They are generally 22 nucleotides long, short segments, and they usually target specific mRNA and silence it. Each one of the miRNA may act upon many mRNAs.

As He and Hannon state:

Non-coding RNAs participate in a surprisingly diverse collection of regulatory events, ranging from copynumber control in bacteria1 to X-chromosome inactivation in mammals2.MicroRNAs (miRNAs) are a family of 21–25-nucleotide small RNAs that, at least for those few that have characterized targets, negatively regulate gene expression at the post-transcriptional leve.

Members of the miRNA family were initially discovered as small temporal RNAs (stRNAs) that regulate developmental transitions in Caenorhabditis elegans6. Over the past few years, it has become clear that stRNAs were the prototypes of a large family of small RNAs, miRNAs, that now claim hundreds of members in worms, flies, plants and mammals.

The functions of miRNAs are not limited to the regulation of developmentally timed events. Instead, they have diverse expression patterns and probably regulate many aspects of development and physiology. Although the mechanisms through which miRNAs regulate their target genes are largely unknown, the finding that at least some miRNAs feed into the RNA INTERFERENCE (RNAi) pathway has provided a starting point in our journey to understand the biological roles of miRNAs.

miRNAs are simple yet complex entities and key players in the epigenetics which control gene expression.

It is clear from the above that miRNAs can positively and negatively impact many elements in the pathways we have considered in HGPIN and PCa. Coppola et al review several of the key ones. For example:

·       miR-146: Down regulates the AR.

·       miR-34: Can recapitulate p53 resulting in apoptosis and arrest.

·       miR-23: can result in c-myc overexpression and cell proliferation.

In a recent paper by Poliseno et al they have identified several others:

·       miR-106b: Down-regulates PTEN and triggers PIN in murine models.

·       miR-22, miR-25, miR-302: Down-regulating of PTEN.

Similarly the papers by Petrocca et al and that by Calin and Croce detail many of the miRNAs and their impacts on many cancers. As seen in the above graphic these are but a few in the overall targeting of PCa control genes. As Coppola et al state:

The hypothesis that miRs can be regarded as new broad-spectrum oncogenes or tumor suppressor genes has opened a revolutionary field of research with exciting diagnostic and therapeutic perspectives.

The compelling hint of a widespread miR deregulation in cancer pathogenesis came from the analysis of the genomic distribution of 186 miR. In this study, it was demonstrated that more than half of them mapped in cancer-associated genomic regions, namely in chromosomal sites prone to deletions, amplifications or recombinations. These aberrations can result in miR down- or up-regulation, conferring selective advantages to mutated cells.

Additional mechanisms of miR deregulation include altered expression of miRs as a consequence of excessive or deficient processing; aberrant transcription of the precursors by epigenetic silencing of miR promoters or as a result of the activity of oncogenic transcription factors; and more rarely, point mutations in mature miRs or in target sequences that can interfere with normal target recruitment

The problem that we will have in any modeling of HGPIN and PCa is not only do we have issues regarding the somewhat well-known genes but the impact of the epigenetic factors is unknown, complex, and possibly random.

Furthermore miRNAs can act in a positive or negative manner depending upon the cell and the activated networks in the cell. From Croce (2009) we have:

Importantly, miRNAs should not be described as oncogenes or tumor suppressor genes, unless the tissue or cell type involved in their action is specified. For example, miR-221 and miR-222 target an oncogene, KIT, and inhibit the growth of erythroblastic leukaemia30, and therefore function as tumor suppressors in erythroblastic cells. but they also target at least four important tumor suppressors — phosphatase and tensin homologue (PTEN), p27, p57 and tissue inhibitor of metalloproteinases 3 (TIMP3) — and function as oncogenic miRNAs by suppressing these tumor suppressors in various human solid tumours31 (TABLE 1). Therefore, before describing an miRNA as a tumor suppressor or an oncogene, it is necessary to specify in which cell or tissue, as cellular context is crucial for the function of miRNAs….

Recent work on miR-34 has demonstrated its impact on p53 (Rokhlin et al) and the fact that miR-34 significantly mediates the role of p53 in apoptosis in AR dependent PCa.

As Sevli et al state:

The miRNAs have critical functions in gene expression and their dysregulation may cause tumor formation and progression. Today, it is known that tumors possess widespread deregulated miRNA levels. Over-expression or down-regulation of specific miRNAs in different tumor types make them potential therapeutic targets and diagnostic markers. Up-regulated miRNAs inhibiting tumor suppressor genes in tumor cells are commonly termed as oncogenic miRNAs or oncomirs. The miRNAs whose down-regulation promotes tumor progression are tumor suppressor miRNAs. One type of mRNA may possibly be targeted by multiple different miRNAs with variable efficiencies. Conversely, a single miRNA may target more than one mRNA. Thus, to be able to observe a tumorigenic phenotype, some significant changes should occur in microRNome content of the cells.

MiRNA and Stem Cells

As we have indicated elsewhere, the concept of the cancer stem cell has received significant attention. There has also been a great deal of work on the area of linking miRNAs and the stem cell model for PCa. In a recent work by Liu et al (2011) the authors demonstrate the nexus between miR-34a and its ability to inhibit PCa stem cells by directly repressing CD44. They observe that cancer stem cells have been observed in many solid cancers by using the fact that CD44 adheres to the cell surface. PCa stem cells with enhance clonogenic and tumor initiating and metastatic capacities are often enriched with CD44+ cell population. The work of Liu et al demonstrated that the administration of miR-34a to PCa cells inhibited PCa metastasis and inhibited PCa regeneration. This is one of the first uses of miRNA as a tumor suppressor.

In a recent paper by Xia (2008) the author states:

The key characteristics of stem cells are that they are capable of self-renewal and differentiation. The mechanisms by which stem cells maintain self-renewal and differentiation are complicated. In the past years, protein-coding genes had been broadly investigated in stem cell self-renewal and differentiation.

Recent studies indicate miRNAs as one of the most abundant classes of post-transcriptional regulators proved to be crucial in a wide range of biological processes, which suggest that miRNAs may also play essential roles in stem cell self-renewal and differentiation. Disruption of Dicer function in murine ESs influences miRNA processing and greatly impairs their ability to differentiate …

Cancer stem cells (CSCs) are the cells within a tumor that possess the capacity to self-renew and to produce the heterogeneous lineages of cancer cells that comprise the tumor. CSCs can thus only be defined experimentally by their ability of self-renewal and tumor propagation.

The implementation of this approach explains the use of alternative terms in the literature, such as “tumor-initiating cells” to describe putative CSCs. …

The identification of growth and differentiation pathways responsible for CSC proliferation and survival will help in the discovery of novel therapeutic targets. Previous studies have shown that many signal pathways may participate in regulating CSC functions, including Wnt/β-catenin, Notch, and Sonic hedgehog homolog (SHH). The canonical Wnt cascade has emerged as a critical regulator of stem cells and activation of Wnt signalling has also been associated with various cancers …

CSC maintenance is dependent on β catenin signaling. Moreover, because Wnt/β-catenin signalling is not essential for normal epidermal homeostasis, such a mechanistic difference may thus be targeted to eliminate CSCs and consequently eradicate squamous cell carcinomas. It is therefore hypothesized that inhibition of Wnt signaling may provide an effective way to reduce the unwanted stem cell renewal which results in cancers.

Inhibition of Wnt signalling may prove to be an effective road to inhibit the uncontrolled cell renewal that drives cancer. Acting as novel and pivotal regulators of protein-encoding genes, miRNAs will have great potential in regulating CSCs’ biological functions by targeting CSCs-related signal pathway molecules.

We have performed various analyses of CSCs especially for PCa. This is a critical area for ongoing research and most likely will prove quite useful.

MET

MET is a tyrosine kinase receptor. It is activated by HGF the hepatic growth factor and it in turn activates a multiplicity of pathways. It is considered a proto-oncogene and thus is of general concern. From NCBI[4]:

The proto-oncogene MET product is the hepatocyte growth factor receptor and encodes tyrosine-kinase activity. The primary single chain precursor protein is post-translationally cleaved to produce the alpha and beta subunits, which are disulfide linked to form the mature receptor. Various mutations in the MET gene are associated with papillary renal carcinoma.

MET is located on 7q31. We now examine the MET structure and then examine its control over several pathways.

From Benvenuti and Comoglio we have:

Both MET and RON are tyrosine kinases crucially involved in the control of the ‘‘invasive growth’’ (Giordano et al., 2002). Under physiological conditions such as embryonic development and organ regeneration, they contribute to establishing the normal tissue patterning by orchestrating cellular proliferation, disruption of intercellular junctions, migration through the EMC and protection from apoptosis. In transformed tissues, receptor deregulation is responsible for cancer progression and metastasis formation and dissemination. Either upon ligand stimulation or receptor constitutive activation, cancerous cells are induced to leave the primary tumor, degrade the basal membrane, move towards different organs and there give rise to metastasis

MET controls many pathway elements in a cell. We show some of them in the Figure below from Dickinson and Duncan. Note the HGF binds to MET and thus it activates a set of pathways facilitating invasion and stopping cell cycle arrest.

The above demonstrates the MET pathway and its relationship to the many other key pathways.

MET can be over expressed and over activated and the result is a malignant growth. Thus MET has the potential for becoming a significant factor in cancer development. From Benvenuti and Comoglio:

It has been extensively demonstrated that when used in a deviant cellular environment and without spatial and temporal regulation, MET exerts a major role in tumor formation and progression. Cells which over-express either MET or HGF are tumorigenic when implanted into nude mice and become extremely metastatic, moreover transgenic mice for either MET or HGF develop metastatic tumors  while, on the contrary, endogenously expressing cancer cells become less aggressive when MET is switched off.

In the above the issue is over expression. The question is; what is driving that over expression? Is it truly an excess production or a loss of control or modification? Is this on a cell by cell basis or is it pandemic? They continue:

Accordingly, it was demonstrated that short hairpin RNA (shRNA) mediated MET knockdown in rabdomyosarcomas (RMS)-derived cell lines greatly affects cell proliferation, survival and invasion. Furthermore in xenograft models of RMS MET silencing produced a dramatic reduction of tumor mass. Similar results were obtained silencing MET in lung cancer cell lines harboring MET amplification. In those cell lines receptor silencing (once more achieved by shRNA technology) induced a significant growth inhibition; notably the silencing sorted no effects on cell lines that did not display receptor gene amplification.

This seems to answer the question regarding complete cell line activation.

It has been extensively described, both in animal models and in normally occurring human cancers, that constitutive activation of MET can be achieved in three different ways:

(i) with establishment of ligand-receptor autocrine loops;

(ii) via receptor over-expression, and

(iii) in presence of activating point mutations in the receptor coding sequence.

Ligand-receptor autocrine circuits make cells independent from the need of growth factors; receptor over-expression triggers receptor oligomerization and reciprocal activation even in absence of ligands; point mutations generate constitutively active receptors.

This last event is extremely uncommon; however, some missense point mutations have been described in MET coding sequence in certain human cancers.

The above discussion describes the ways in some detail. The causes of over expression could then be addressed as a therapeutic methodology. They continue:

Particularly missense mutations located in the tyrosine kinase domain of MET were described in patients who suffer from hereditary and sporadic papillary renal-cell carcinomas and head and neck squamous-cell carcinomas, whereas alterations in the juxta-membrane region were mainly found in human gastric and lung cancers

The above does also present the issue of mis-sense mutations, changes that may not change anything but may cause a cessation of genetic progression.

Observations

The paper which we have used to initialize the focus on this report is one which combines: mir-34, MET, p53, and methylation. It is an amalgam of receptors of the kinase inhibitor variety, key pathway oncogenes, miRNAs and methylation. It is an interplay between all of the complex elements which are now known in cancer genetics.

The Cheng et al results are simply as follows:

1.     miR-34 Cooperates with p53 in Suppression of Prostate Carcinogenesis

2.     p53 and miR-34 Cooperate in the Control of Prostate Stem/Progenitor Cell Activity

3.     p53 and miR-34 Regulation of Stem/Progenitor Cells Depends on MET

However in their conclusions we have also introduced the methylation effects as well. They conclude:

Our study provides direct genetic proof that miRNAs of the miR-34 family may act as tumor suppressors in concert with other genes, such as p53. These findings offer a solid physiological basis for the rational design of diagnostic and therapeutic approaches. Because the lack of mir-34 genes alone is insufficient for cancer initiation, their downregulation is likely to occur at some point during tumor progression.

However, the preexistence of mir-34 methylation in some normal cells cannot be excluded. Further genomic studies in conjunction with animal modeling should be able to address this question. Although our current studies have been focused on prostate cancer, tissue-specific inactivation of mir-34 and p53 in other tissues will address likely interactions of these genes in other cell lineages.

Thus we have exhibited here a complex interplay between types of cell control mechanisms. The challenge will be how best to model this complex interplay. In our prior analyses we have let epigenetic factors be secondary and considered almost as noise. Here, however, they are pari passu with all other elements and must be considered expressly.

Also Liu et al from Tang’s Lab state:

We have shown that miR-34a is underexpressed in tumorigenic CD44+ prostate cancer cells and that it has potent antitumor and antimetastasis effects. Our results establish miR-34a as a key negative regulator of CD44+ prostate cancer cells and CD44 as an important target of miR-34a. Our findings suggest that reduced expression of miR-34a in prostate CSCs contributes to prostate cancer development and metastasis by regulating expression of CD44 and the migratory, invasive and metastatic properties of CSCs

Tang’s Lab has done extensive work on PCa CSC and the implications of reduced miR-34 are significant. The issue here is several fold. First, the measure of miR-34 activity can be prognostic. Second, the reasons for reduced miR-34 is of prime concern. As we shall note later, the cause may be methylation of CpG clusters. Thus if one were to try anti-methylation drugs, would that assist? There is always a concern here since anti-methylation therapeutics are non-selective.

References:

1.     Abba, M., et al, MicroRNAs in the Regulation of MMPs and Metastasis, Cancers 2014, 6, 625-645; doi:10.3390/cancers6020625.

2.     Araud, T., Overview of the miRNA Pathways, Thesis, U Geneva, 2008.

3.     Armstrong, L., Epigenetics, Garland (New York) 2014.

4.     Bader, A. et al, Developing therapeutic microRNAs for cancer, Gene Therapy, 2011, V 18.

5.     Bartel, D., MicroRNAs: Genomics, Biogenesis, Mechanism, and Function, Cell 2004 pp 281-297.

6.     Benvenuti, S., P., Comoglio, The MET Receptor Tyrosine Kinase in Invasion and Metastasis, J. Cell. Physiol. 213: 316–325, 2007

7.     Calin, G., C. Croce, MicroRNA Signatures in Human Cancers, Nat Rev 2006 pp 857-866.Calin, G., C. Croce, Chromosomal Rearrangements and microRNAs, Jrl Clin Inves 2007 pp 2059-2066.

8.     Cheng, C. Y., et al, miR-34 Cooperates with p53 in Suppression of Prostate Cancer by Joint Regulation of Stem Cell Compartment, Cell Reports (2014), http://dx.doi.org/10.1016/j.celrep.2014.02.023 

9.     Coppola, V., et al, MicroRNAs and Prostate Cancer, Endo Rel Canc 2010 pp F1-F17.

10.  Croce, C., Causes and Consequences of microRNA Dysregulation in Cancer, Nature 2009 pp 704-714.

11.  Gottwein, E., B. Cullen, Viral and Cellular MicroRNAs as Determinants of Viral Pathogenesis and Immunity, Cell Host 2008, pp 375-387.

12.  He, L., G. Hannon, MicroRNAs: Small RNAs with a Big Role in Gene Regulation, Nature, 2004 pp 522-532.

13.  Huang, C., et al, A Study of Cancer Related MicroRNAs through Expression Data and Literature Search, World Acad Sci & Engr, 2009 254-256.

14.  Jones, P., S. Baylin, The Epigenomics of Cancer, Cell, 2007 pp 683-692.

15.  Kasinski, A., F., Slack, miRNA-34 Prevents Cancer Initiation and Progression in a Therapeutically Resistant K-ras and p53-Induced Mouse  model of Lung Adenocarcinoma, Cancer Res Published OnlineFirst September 10, 2012.

16.  Kim, N., et al, A p53/miRNA-34 axis regulates Snail1-dependent cancer cell epithelial–mesenchymal transition, J. Cell Biol. Vol. 195 No. 3 417–433, 2011.

17.  Lawrie, C., MicroRNAs in Medicine, Wiley (Hoboken, NJ) 2014.

18.  Liu et al, The microRNA miR-34a inhibits prostate cancer stem cells and metastasis by directly repressing CD44, Nature Medicine,    17,     211–215     (2011)

19.  Liu, C., et al, The microRNA miR-34a inhibits prostate cancer stem cells and metastasis by directly repressing CD44, nature medicine VOLUME 17, NUMBER 2, FEBRUARY 2011

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22.  Pekarik, V., et al, Prostate Cancer, miRNAs, Metallothioneins and Resistance to Cytostatic Drugs, Current Medicinal Chemistry, 2013, Vol. 20, No. 4

23.  Peruzzi, B., D. Battero, Targeting the c-Met Signaling Pathway in Cancer, Clin Cancer Res 2006;12:3657-3660.

24.  Petrocca, F., et al, Emerging Role of miR-106b-25/miR-17-92 Clusters in the Control of Transforming Growth Factor β Signalling, Can Res 2008, pp 8191-8194.

25.  Poliseno, L., et al, Identification of the miR-106b-25 MicroRNA Cluster as a Proto-oncogenic PTEN-targeting Intron That Cooperates with Host Gene MCM7 in Transformation, Sci Sig 2010 .

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[1] http://www.biosciencetechnology.com/news/2014/03/gene-family-proven-suppress-prostate-cancer also http://www.news.cornell.edu/stories/2014/03/gene-family-proven-suppress-prostate-cancer

[2] http://staging-www.nature.com/nm/journal/v17/n2/full/nm.2284.html

[3] http://www.ncbi.nlm.nih.gov/gene/407040

[4] http://www.ncbi.nlm.nih.gov/gene/4233