“Wars and elections are both too big and too small to matter in the
long run. The daily work—that goes on, it adds up. It goes into
the ground, into crops, into children’s bellies and their bright
eyes. Good things don’t get lost.
Here's what I've decided: the very least you can do in your life is to
figure out what you hope for. And the most you can do is live
inside that hope. Not admire it from a distance but live right inside
it, under its roof. What I want is so simple I almost can't say it:
elementary kindness. Enough to eat, enough to go around. The
possibility that kids might one day grow up to be neither the
destroyers nor the destroyed. That's about it. Right now I'm living
in that hope, running down its hallway and touching the walls on
both sides. I can't tell you how good it feels”
From Barbara Kingsolver, “Animal Dreams”
Despite Koch’s discovery, for 60
years the only treatment was
isolation, “fresh air, and sunshine”
www.umdnj.edu/librweb
Waverly Hills Tuberculosis Sanatorium. Louisville KT 1926
Built in 1911, expanded in 1926, closed in 1961
So it was time for another Nobel laureate!
Selman Waksman
1888-1973
He was working on bugs that live in dirt!
Selman Waksman
1888-1973
He was working on bugs that live in dirt!
A childhood immigrant from the
Ukraine, through hard work
He became a professor
Of microbiology and biochemistry
at Rutgers
Selman Waksman
1888-1973
He was working on bugs that live in dirt!
A childhood immigrant from the
Ukraine, through hard work
He became a professor
Of microbiology and biochemistry
at Rutgers
He coined the term antibiotic and
Discovered more than 20
Antibiotics, including two which
Are widely used today.
Neomycin--check your medicine cabinet
Selman Waksman
1888-1973
He was working on bugs that live in dirt!
Streptomyces griseus.
Streptomycin
He was working on bugs that live in dirt!
Streptomyces griseus.
Streptomycin
Isolated on October 19, 1943 by Albert Schatz,
a graduate student in Waksman’s lab
He was working on bugs that live in dirt!
the-scientist.com
Streptomycin
Isolated on October 19, 1943 by Albert Schatz,
a graduate student in Waksman’s lab
Bugs fight bugs
Streptomycin kills many bacteria, including
Mycobacterium Tuberculosis!
Bugs fight bugs
Streptomycin kills many bacteria, including
Mycobacterium Tuberculosis!
But how?
The central dogma-anybody remember that?
The central dogma-anybody remember that?
DNA -> RNA -> Proteins
The central dogma-anybody remember that?
DNA -> RNA -> Proteins
And we call that step?
DNA -> RNA -> Proteins
Translation!
DNA -> RNA -> Proteins
Bacteria do it too,
and like us they use RIBOSOMES
www.palaeos.com
Let’s zoom in on the action
Lovely picture from Harry Noller
The ribosome is an amazing machine
That unlike most in the cell runs on RNA!
Lovely picture from Harry Noller
The ribosome is an amazing machine
That unlike most in the cell runs on RNA!
16S rRNA + proteins =30S or small subunit
Lovely picture from Harry Noller
If we zoom in further…
Luckily, our ribosomes are slightly different
and thus streptomycin affects them less
16S rRNA + proteins =30S subunit
Lovely picture from Harry Noller
Streptomycin cannot be given orally, but must be
administered by regular intramuscular injection.
An adverse effect of this drug is ototoxicity, i.e.
It can result in temporary hearing loss.
Streptomycin cannot be given orally, but must be
administered by regular intramuscular injection.
An adverse effect of this drug is ototoxicity, i.e.
It can result in temporary hearing loss.
Cool fact--this may be
due to effect on
Mitochondrial ribosomes!!!
OK. So streptomycin kills bugs in
a flask in the lab. What about
inside a patient?
First we have to make a lot of it.
In steps George Merck
of Merck and Co.
First we have to make a lot of it.
In steps George Merck
of Merck and Co.
And we got the patent…
First we have to make a lot of it.
In steps George Merck
of Merck and Co.
But we gave it back to Rutgers!
Now we need to try it on animals.
Enter Dr. William H. Feldman and Dr. H. Corwin Hinshaw
at the Mayo Clinic
Now we need to try it on animals.
Enter Dr. William H. Feldman and Dr. H. Corwin Hinshaw
at the Mayo Clinic
In two months they reported to Waksman
that four tubercular guinea pigs receiving streptomycin
"look exceedingly well."
We do, don’t we!
OK, but how about people?
Next Feldman and Hinshaw invent clinical trials
www.jameslindlibrary.org
OK, but how about people?
Next Feldman and Hinshaw invent clinical trials
OK, but how about people?
Next Feldman and Hinshaw invent clinical trials
This is really important!!
OK, but how about people?
In August 1945 Hinshaw reported that
thirty-three patients had been treated
"and [we] continue to be quite optimistic."
Initially streptomycin
appeared to be a miracle cure
Patients, including the first, “Patricia T”,
Were returned to health from death’s door
Now the problem was scaling up
(Remember 5 million people a year
Were still dying of TB!)
By 1948 8 companies were making streptomycin
But their 80,000 pounds
Would only treat 1000 patients
Initially streptomycin
appeared to be a miracle cure
BUT…..
A second, much worse
Problem was now on the horizon
By 1948 patients began to relapse!
For example the author George Orwell
But a second, much worse
problem was now on the horizon
In an MRC clinical trial, patients improved rapidly
But within five years the death rate was the
Same as the untreated controls
Professor Hill of the MRC
But a second, much worse
problem was now on the horizon
In an MRC clinical trial, patients improved rapidly
But within five years the death rate was the
Same as the untreated controls
Uh, oh.
Professor Hill of the MRC
What was going wrong??
What was going wrong??
Streptomycin resistant bacteria
could be cultured from these patients!
What was going wrong??
Streptomycin resistant bacteria
could be cultured from these patients!
How could
That happen?
What was going wrong??
Streptomycin resistant bacteria
could be cultured from these patients!
How could
That happen?
Have you heard
The one about
Natural selection?
What was going wrong??
evolution.berkeley.edu
And how did this happen?
evolution.berkeley.edu
And how did this happen?
evolution.berkeley.edu
And how did this happen?
evolution.berkeley.edu
And how did this happen?
evolution.berkeley.edu
And how did this happen?
Remember me?
16S rRNA + proteins =30S subunit
Lovely picture from Harry Noller
Occasional bacteria had variants in
16sRNA or the ribosomal protein S12
16S rRNA + proteins =30S subunit
Lovely picture from Harry Noller
These blocked binding of streptomycin
to the ribosome and thus these bacteria
were resistant!
16S rRNA + proteins =30S subunit
Lovely picture from Harry Noller
Thanks and kudos to Gary Trudeau for hitting the nail on the head
So what now?
So what now?
Luckily, help
was on the way
from
para-aminosalycilic acid (PAS)
AND
Jörgen Lehmann
Lehmann started with a strange published fact
The TB bug loves to eat aspirin (salicylic acid)
Lehmann started with a strange published fact
The TB bug loves to eat aspirin (salicylic acid)
Lehmann started with a strange published fact
The TB bug loves to eat aspirin (salicylic acid)
His hypothesis: Alter aspirin slightly
And the bugs will die trying to eat it
Lehmann moved quickly,
from bacterial trials in December 1943
To Guinea pig trials in January 1944
To the first patient in March 1944
(before streptomycin!!)
Lehmann J. Para-aminosalicylic acid
in the treatment of tuberculosis. Lancet. 1946; 1:15-6.
Chest 1949;16;684-703
So how does PAS work?
So how does PAS work?
50 years later we
still don’t know for sure
So how does PAS work?
It is likely to act somewhere in the pathway
For making nucleotides to make DNA
http://themedicalbiochemistrypage.org
So which is better:
Streptomycin or PAS?
So which is better:
Streptomycin or PAS?
BOTH!
Remember this?
I fear no streptomycin
Remember this?
I fear no streptomycin
and PAS doesn’t touch me
But if the same patient is given both drugs…
I fear no streptomycin
and PAS doesn’t touch me
But if the same patient is given both drugs…
Ahhhh……
No………
After this trial…..
The two drug combination became
The new standard
But the story is not over yet-- we needed
Another Nobel laureate
But the story is not over yet-- we needed
Another Nobel laureate
I was already the 1939 Nobel laureate
For discovering sulphanilamide
The first antibacterial drug
Gerhard Domagk
Nobelprize.org
But the story is not over yet-- we needed
Another Nobel laureate
I was already the 1939 Nobel laureate
For discovering sulphanilamide
The first antibacterial drug
(though I was not allowed by the Nazis
to accept the prize and I was held
by the Gestapo for a week
because of this honor)
Gerhard Domagk
Nobelprize.org
But the story is not over yet-- we needed
Another Nobel laureate
I spent World War II in a bombdamaged hospital lab
pursuing new chemical relatives of
sulphanilamide that
might be effective against TB
Gerhard Domagk
Nobelprize.org
Here’s what I discovered
Isoniazid
Here’s what I discovered
First we need to know more
About mycobacteria
Isoniazid
All living cells
Have a plasma
membrane
All living cells
Have a plasma
membrane
Gram positive
Bacteria also
Have a cell wall
All living cells
Have a plasma
membrane
Gram positive
Bacteria also
Have a cell wall
Mycobacteria have a
Very complex cell wall
Serving as a barrier
http://web.uct.ac.za/depts/mmi/lsteyn/cellwall.html
1.
2.
3.
4.
5.
6.
7.
8.
outer lipids
mycolic acid
polysaccharides (arabinogalactan)
peptideglycan
plasma membrane
lipoarabinomannan (LAM)
phosphatidylinositol mannoside
cell wall skeleton
Wikipeda/mycobacterium
Just look at
the parts list
1.
2.
3.
4.
5.
6.
7.
8.
outer lipids
mycolic acid
polysaccharides (arabinogalactan)
peptideglycan
plasma membrane
lipoarabinomannan (LAM)
phosphatidylinositol mannoside
cell wall skeleton
Wikipeda/mycobacterium
Just look at
the parts list
Isoniazid is metabolized by the bacterium
To isonicotinic acyl anion or radical
and inhibits the enzyme
that makes
Isoniazid was added to the combination therapy
and remains there today!
Isoniazid was added to the combination therapy
and remains there today!
Streptomycin and PAS were removed from
frontline therapy as new drugs
that were more effective or
had fewer side effects
came into the clinic
Current first-line TB drugs
and their dates of discovery
National Institute of Allergy and Infectious Diseases (NIAID) 2007
NIAID (2007)
Let’s look at one of the new drugs in detail
rifampicin
It binds the ß subunit
of RNA polymerase
rifampicin
Cell. 2001 Mar 23;104(6):901-12
Check out a cool video view at
http://www.pingrysmartteam.com/rifampicin/rifampicin.htm
Binding does not block recruitment to the promotor
But inhibits transcription elongation after 2-3 nucleotides are made
by sterically blocking transcript elongation
Cell. 2001 Mar 23;104(6):901-12
Let’s take a closer look
Cell. 2001 Mar 23;104(6):901-12
Most mutations that confer rifampicin resistance change
Residues in the binding pocket
Our RNA polymerase is different enough that it is not
inhibited by rifampicin
Cell. 2001 Mar 23;104(6):901-12
First-line TB drugs
Left to right isoniazid, rifampin, pyrazinamide, & ethambutol
Streptomycin (not shown) is given by injection
CDC
First-line TB drugs
4 drugs, 130 doses
Over 6-9 months!
TBAlliance
First-line TB drugs
4 drugs, 130 doses!
Daily dose!
http://www.mcvay.com/tb_treatment_in_the_western.html
TBAlliance
First-line TB drugs
Are also not compatible with
Antiretroviral HIV therapy
(problem for 1/3 of HIV patients!)
Rifampin induces cytochrome p450s,
Increasing metabolism of anti-HIV drugs
TBAlliance
Remember the difference
in TB rates world-wide but
CDC
The other major challenge
Is multi-drug resistant TB
CDC
Remember this story???
In other word MDR TB says
I fear none of the
First line drugs!
What drives drug resistance?
Incorrect, unsupervised, or incomplete treatment
The answer in the developed world:
The answer in the developed world:
MORE DRUGS!
NIAID (2007)
NIAID (2007)
New TB drugs under development
NIAID (2007)
And the future beckons
Nature 393, 537-544(11 June 1998)
In 1998 the Sanger centre and the Pasteur Institute
Determined the complete genome sequence
Of M. tuberculosis
Nature 393, 537-544(11 June 1998)
4.4 million base pairs
About 4000 genes
Remarkable diversity
Of genes involved in fatty acid
Synthesis and metabolism
(>250 genes)
Nature 393, 537-544(11 June 1998)
Remarkable diversity
Of genes involved
in fatty acid
Synthesis and
metabolism
(>250 genes)
Nature 393, 537-544(11 June 1998)
The genome provides
new targets for
drugs and vaccines
Nature 393, 537-544(11 June 1998)
But in the developing world…..
Stats from WHO; slide from TBAlliance