International MPN News, Science & Opinion

The Silent Gene

The Silent Gene — Panobinostatpanobinostat formula

by Zhenya Senyak

There’s a thin, six foot ribbon of DNA in the nucleus of our cells.

On it is written a story more than 3.5 billion years old.  The last chapter is our personal story – uniquely our own – and leading up to it is the story of life on earth from virus and protozoan on up through fish, reptile and all the rest that came before us.

The identical story is repeated trillions of times in the nucleus of every cell.  Cells of the human eyes, nerve cells, muscle cells, blood and bone cells all so different in function have the exact same story written in their DNA.

(If you prefer pictures to tell this part of the story there is a neat Nova slide show here.)

It’s all pretty incomprehensible, impossible to grasp visually, but a simple question casts light on a major research achievement affecting everyone with DNA

Q:  How do you tuck a six foot strand of 3.5 billion year old DNA history inside the nucleus of a cell that is invisible to the naked eye?

A:  You wind it very tightly around a spool.

Why this matters to us as patients has to do with the way counter-intelligence war is waged against MPNs.

The major MPN therapeutic  thrust of the past few years has been inhibition.  Once we cracked the code used by enzymes – JAK2, in one instance – to signal production of blood lines, we found ways to break up the communication, inhibit the action.  Enter Jakafi, for example.

Since the JAK-STAT pathway along which this signal runs also signals many other biological events, our inhibition stopped other processes – some good, some bad.  Without understanding exactly the full mechanism of action, we could often get a good result, for a time.

But suppose instead of intercepting the code and inhibiting it we prevent the message from being issued in the first place? We silence the gene signalling the action.

Which takes us back to the tightly wrapped DNA strand.  Actually, it’s VERY tightly wrapped DNA in highly specific patterns and bonded by its negative electrical charge to aNucleosome1 positively charged spool called a histone. 

We know that genes – small stretches of DNA found on one or another of our 23 chromosomes – can be turned on, expressed, by events in the body to initiate a cascade of events that can be transcribed to produce proteins to defend or sustain the body.  Sometimes, due to genetic modification or mutation, one or more genes can get a little out of hand and produce excessive  proteins that are destructive to the body.  Too many platelets,  too many red blood cells.

For genes to be turned on, the DNA strand of which it is a part has to loosen up a bit from the histone around which it’s wrapped.  Otherwise the gene would not be accessible for transcription.

The process of loosening that strand — sort of like opening a book to a page so it can be read —  is started by introducing an acetyl molecule into the area and so it’s called acetylation (Ah-SEE-till-a-shun)  or lysine acetylation lysinesince acetylation occurs on the residue of lysine, an essential amino acid.

Histone deacetylase (HDAC) enzymes help silence the gene by preventing acetylation  or de-acetylizing the gene — and keeping the DNA strand electronically glued to its histone spool.

So now let’s leap to the bottom line to see how this affects a patient with myelofibrosis.

Panobinstat is an experimental HDAC used in clinical trial run by, among others, Dr. John Mascarenhas and Dr. Ronald Hoffman, both of Mt. Sinai hospital.  The drug was developed and manufactured by Novartis which funded  and collaborated in the trial.

Before After Trichrome Stain

This is what a successful outcome might look like.

 

Published this month, the paper in the British Journal of Haematology by John Mascarenhas et al. includes this photograph of comparative slides made from processing a bone marrow specimen from a single patient (see Alice, below) after 16 cycles of Panobinostat.  The slide, reads the paper’s caption:  “..shows hypercellular marrow with increases in all haematopoietic lineages….There is partial resolution of fibrosis with only moderately increased reticulin fibres and minimal collagen deposition.”

Dr. Attilio Orazi Weill-Cornell

Dr. Attilio Orazi
Weill-Cornell

Although it looked pretty clear to our non-professional eyes that the photo on the right was clearer of fibrosis and looked a lot like normal bone marrow, MPNforum submitted the photograph to Dr. Attilio Orazi, the chief hematopathologist at Weill-Cornell and a member of the MPNclinic, for an independent assessment.  Although he noted he needed a better image to do a full evaluation, he concurred:  “The marrow seems more cellular with less collagen by trichrome stain….”

More cellular, in our terms, means more like normal blood cell lines and less collagen is reduced fibrosis. (Trichrome  is a three color staining process to permit viewing cells apart from their  environment. )

Dr. Ron Hoffman

Dr. Ron Hoffman

Commenting in the Mount Sinai blog VISION on the need for long-term, low dose application of Panobinostat (LBH589) to achieve results, Mascarenhas recalled “Persistence led us to finally see signals of activity….One of our patients came in for a checkup and her blood smear looked normal. I handed it to Dr. (Ron) Hoffman to look at and asked him what he thought was wrong with the patient. He said “Nothing. It’s normal.’ Then I told him it was one of our patients in the trial.”

The Panobinostat trial itself was so small  — 18 patients reduced to five after toxicity forced the others to drop out — and so extended — over three years — that it attracted only slight interest when results were first pubished in 2009.  And then again, with updated results,  at ASH, in 2011.  Publication of Mascarenhas et al. paper in the BJH last month — and the  front page story in the Mt. Sinai blog — refocused attention on the finding in the MPN patient community.  Suddenly we were asking what’s Panibinostat and how do I sign up for the clinical trial?

The clinical trial — then and now 

Panobinostat (LBH589) is a Novartis Pharmaceuticals drug. So is ruxolitinib (Jakafi).  The joining of these two drugs in clinical trial has been long in development.

From the beginning Novartis has worked both collaboratively and independently with researchers on Panobinostat clinical trials. The first safety and efficacy Novartis Panobinostat trial for patients with MF was in 2009. A clinical Panobinostat trial

Dr. John Mascarenhas Mt. Sinai

Dr. John Mascarenhas
Mt. Sinai

referenced by Dr. John Mascarenhas in BJH was reported at ASH in 2009 and completed in 2011. That year at the ASH San Diego meeting,  Novartis employees Fabienne Baffert, et al. presented a paper on mouse studies demonstrating improved efficacy using ruxolitinib with panobinostat to impact mutant JAK2 pathways (JAK2 v617F).

The phase one 18 patient study (Clinical trial NCTO 1298934) sponsored by Dr. Ronald Hoffman (Mt. Sinai School of Medicine), with the collaboration of Novartis, led directly to the new series of panobinostat/ruxolitinib combinatorial trials now recruiting patients.

At first, you might not think it was so promising.  The initial group of 20 recruits was whittled down to 18 after screening.  This was a dose limiting toxicity trial, so as patients’ counts went down — thrombocythemia (platelets) — were the first causes for drop out and then anemia.  Almost all had some adverse events like diarrhea but no serious adverse events that would have called into question the safety of the drug.  In the end there were only five remaining patients and, for the history books, only one — we’re calling her Alice so we can refer to people instead of numbers — who was a special case.

Here’s Alice

Even though toxicity limited continued participation of most patients in the trial, the adjustment of dosage  — smaller doses over longer periods of time — seemed to deal with the worst of the problems. “Panobinostat was effective in improving MF-related symptons in the majority of patients treated for longer than a month,” wrote Mascarenha. “Significant reduction in splenomegaly was observed at the end of six cycles…Importantly a near CR (complete response) was obtained in a single patient after >12 cycles of  therapy at reduced dos(age)…”

That single patient was Alice.  And of the remaining patients still on trial and evaluable for response after six cycles of panobinostat, Alice is a standout.  She was the only patient whose MF was secondary to essential thrombocythemia, the least aggressive chronic MPN.  And she was one of two who did not have the JAK2 mutation.

Most importantly, she did have a complete response (look at the slides), was no longer transfusion dependent, and like the other patients evaluable after six cycles of the drug, her spleen was no longer palpable.

The Hoffman Mt. Sinai trial, despite its small size and high drop-out rate due to toxicity, was successful enough to warrant  a long extension and help power the new combinatorial trial sponsored by his Mt. Sinai associate John Mascarenhas (Clinical trial NCTO 1693601).  The unmistakable evidence of complete response, fibrosis reversal and restored hematopoietic function is a powerful argument.  Linking Panobinostat with the proven track record and limitations of Jakafi — which also has issues with thrombocyopenia and anemia — does not at first seem to make sense.  But the whole is greater than the sum of the parts and, as suggested in the Baffert et al paper, above,  there is some evidence of synergy between the two drugs that could prove powerful.

Novartis thinks so.  The giant pharma is fielding its own very similar safety and dosage clinical trial  of Panobinostat and Ruxolitinib (Clinical Trial: NCTO 1433445) at locations throughout Europe.

Take me back to the Contents

© Zhenya Senyak and MPNforum.com, 2013. Unauthorized use and/or duplication of this material without express and written permission is strictly prohibited. Excerpts and links may be used, provided that full and clear credit is given to MPNforum.com with appropriate and specific direction to the original content.

Comments on: "The Silent Gene" (8)

  1. Robert Baumeister said:

    There is an update on the dose finding study on the combination of Ruxolitinib and Panobinostat from ASH 2013. Quite impressive preliminary data in terms of spleen size reduction. About 50% of all participants in the study achieved a 100% (!) reduction in spleen size. All of the patients in the cohort six with the highest dose of the combination achieved a 100% spleen size. As mentioned in the above article there are some issues with toxisity of the Panobinostat, mainly GI (nausea, diharrea). However, the combination is described as “well tolerated”.

    Here is the link to the full presentation. It requires a login.

    http://www.primeoncology.org/online_education/heme/2013/neworleans_vp_exus.aspx

  2. Mary Morochnick said:

    Zhen, Thanks for making the confusing so much more understanding. Those slides are impressive. Wish we could find an Alice for each of us. Some day maybe.

  3. Dave Dickover said:

    Thanks!! Any trials in the states?

    Dave

  4. When a confusing complex is explained simply and clearly while making it entertaining the writer knows something about it. Lucky for us Zhen is among that number.in his beautifully clear & cogent story about Alice in wonderland among Pano and Ruxo. A.

    • Thanks, Arch…but the truth be told I didn’t know a histone from a baritone before working on this article.(I just thought of it as an enzyme.) One of the unalloyed joys of my job is research. Imagine discovering our DNA is electrically welded to a moon rock looking substance

  5. Thankful for the research!

  6. Exciting news!

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