The central therapeutic challenge of the newest cancer medicine, then, was to find, among the vast numbers of similarities in normal cells and cancer cells, subtle differences in genes, pathways, and acquired capabilities--and to drive a poisoned stake into that new heel.
It was one thing to identify an Achilles' heel--and quite another to discover a weapon that would strike it. Until the late 1980s, no drug had reversed an oncogene's activation or a tumor suppressor's inactivation. Even tamoxifen, the most specific cancer-targeted drug discovered to that date, works by attacking the dependence of certain breast cancer cells on estrogen, and not by directly inactivating an oncogene or oncogene-activated pathway. In 1986, the discovery of the first oncogene-targeted drug would thus instantly galvanize cancer medicine. Although found largely serendipitously, the mere existence of such a molecule would set the stage for the vast drug-hunting efforts of the next decade.
The disease that stood at the pivotal crossroads of oncology was yet another rare variant of leukemia called acute promyelocytic leukemia--APL. First identified as a distinct form of adult leukemia in the 1950s, the disease has a distinct characteristic: the cells in this form of cancer do not merely divide rapidly, they are also strikingly frozen in immature development. Normal white blood cells developing in the bone marrow undergo a series of maturational steps to develop into fully functional adult cells. One such intermediate cell is termed a promyelocyte, an adolescent cell on the verge of becoming functionally mature. APL is characterized by the malignant proliferation of these immature promyelocytes. Normal promyelocytes are loaded with toxic enzymes and granules that are usually released by adult white blood cells to kill viruses, bacteria, and parasites. In promyelocytic leukemia, the blood fills up with these toxin-loaded promyelocytes. Moody, mercurial, and jumpy, the cells of APL can release their poisonous granules on a whim--precipitating massive bleeding or simulating a septic reaction in the body. In APL, the pathological proliferation of cancer thus comes with a fiery twist. Most cancers contain cells that refuse to stop growing. In APL, the cancer cells also refuse to grow up.
Since the early 1970s, this maturation arrest of APL cells had prompted scientists to hunt for a chemical that might force these cells to mature. Scores of drugs had been tested on APL cells in test tubes, and only one had stood out--retinoic acid, an oxidized form of vitamin A. But retinoic acid, researchers had found, was a vexingly unreliable reagent. One batch of the acid might mature APL cells, while another batch of the same chemical might fail. Frustrated by these flickering, unfathomable responses, biologists and chemists had turned away after their initial enthusiasm for the maturation chemical.
In the summer of 1985, a team of leukemia researchers from China traveled to France to meet Laurent Degos, a hematologist at Saint Louis Hospital in Paris with a long-standing interest in APL. The Chinese team, led by Zhen Yi Wang, was also treating APL patients, at Ruijin Hospital, a busy, urban clinical center in Shanghai, China. Both Degos and Wang had tried standard chemotherapy agents--drugs that target rapidly growing cells--to promote remissions in APL patients, but the results had been dismal. Wang and Degos spoke of the need for a new strategy to attack this whimsical, lethal disease, and they kept circling back to the peculiar immaturity of APL cells and to the lapsed search for a maturation agent for the disease.
Retinoic acid, Wang and Degos knew, comes in two closely related molecular forms, called cis-retinoic acid and trans-retinoic acid. The two forms are compositionally identical, but possess a slight difference in their molecular structure, and they behave very differently in molecular reactions. (Cis-retinoic acid and trans-retinoic acid have the same atoms, but the atoms are arranged differently in the two chemicals.) Of the two forms, cis-retinoic acid had been the most intensively tested, and it had produced the flickering, transient responses. But Wang and Degos wondered if trans-retinoic acid was the true maturation agent. Had the unreliable responses in the old experiments been due to a low and variable amount of the trans-retinoic form present in every batch of retinoic acid?
Wang, who had studied at a French Jesuit school in Shanghai, spoke a lilting, heavily accented French. Linguistic and geographic barriers breached, the two hematologists outlined an international collaboration. Wang knew of a pharmaceutical factory outside Shanghai that could produce pure trans-retinoic acid--without the admixture of cis-retinoic acid. He would test the drug on APL patients at the Ruijin Hospital. Degos's team in Paris would follow after the initial round of testing in China and further validate the strategy on French APL patients.
Wang launched his trial in 1986 with twenty-four patients. Twenty-three experienced a dazzling response. Leukemic promyelocytes in the blood underwent a brisk maturation into white blood cells. "The nucleus became larger," Wang wrote, "and fewer primary granules were observed in the cytoplasm. On the fourth day of culture, these cells gave rise to myelocytes containing specific, or secondary, granules . . . [indicating the development of] fully mature granulocytes."
Then something even more unexpected occurred: having fully matured, the cancer cells began to die out. In some patients, the differentiation and death erupted so volcanically that the bone marrow swelled up with differentiated promyelocytes and then emptied slowly over weeks as the cancer cells matured and underwent an accelerated cycle of death. The sudden maturation of cancer cells produced a short-lived metabolic disarray, which was controlled with medicines, but the only other side effects of trans-retinoic acid were dryness of lips and mouth and an occasional rash. The remissions produced by trans-retinoic acid lasted weeks and often months.
Acute promyelocytic leukemia still relapsed, typically about three to four months after treatment with trans-retinoic acid. The Paris and Shanghai teams next combined standard chemotherapy drugs with trans-retinoic acid--a cocktail of old and new drugs--and remissions were prolonged by several additional months. In about three-fourths of the patients, the leukemia remission began to stretch into a full year, then into five years. By 1993, Wang and Degos concluded that 75 percent of their patients treated with the combination of trans-retinoic acid and standard chemotherapy would never relapse--a percentage unheard of in the history of APL.
Cancer biologists would need another decade to explain the startling Ruijin responses at a molecular level. The key to the explanation lay in the elegant studies performed by Janet Rowley, the Chicago cytologist. In 1984, Rowley had identified a unique translocation in the chromosomes of APL cells--a fragment of a gene from chromosome fifteen fused with a fragment of a gene from chromosome seventeen. This created an activated "chimeric" oncogene that drove the proliferation of promyelocytes and blocked their maturation, thus creating the peculiar syndrome of APL.
In 1990, a full four years after Wang's clinical trial in Shanghai, this culprit oncogene was isolated by independent teams of scientists from France, Italy, and America. The APL oncogene, scientists found, encodes a protein that is tightly bound by trans-retinoic acid. This binding immediately extinguishes the oncogene's signal in APL cells, thereby explaining the rapid, powerful remissions observed in Shanghai.
The Ruijin discovery was remarkable: trans-retinoic acid represented the long-sought fantasy of molecular oncology--an oncogene-targeted cancer drug. But the discovery was a fantasy lived backward. Wang and Degos had first stumbled on trans-retinoic acid through inspired guesswork--and only later discovered that the molecule could directly target an oncogene.
But was it possible to make the converse journey--starting from oncogene and going to drug? Indeed, Robert Weinberg's lab in Boston had already begun that converse journey, although Weinberg himself was largely oblivious of it.
By the early 1980s, Weinberg's lab had perfected a technique to isolate cancer-causing genes directly out of cancer cells. Using Weinberg's technique, researchers had isolated dozens of new oncogenes from cancer cells. In 1982, a postdoctoral scientist from Bombay working in Weinberg's lab, Lakshmi Charon Padhy, reported the isolation of yet another such oncogene from
a rat tumor called a neuroblastoma. Weinberg christened the gene neu, naming it after the type of cancer that harbored this gene.
Neu was added to the growing list of oncogenes, but it was an anomaly. Cells are bounded by a thin membrane of lipids and proteins that acts as an oily barrier against the entry of many drugs. Most oncogenes discovered thus far, such as ras and myc, are sequestered inside the cell (ras is bound to the cell membrane but faces into the cell), making them inaccessible to drugs that cannot penetrate the cell membrane. The product of the neu gene, in contrast, was a novel protein, not hidden deep inside the cell, but tethered to the cell membrane with a large fragment that hung outside, freely accessible to any drug.
Lakshmi Charon Padhy even had a "drug" to test. In 1981, while isolating his gene, he had created an antibody against the new neu protein. Antibodies are molecules designed to bind to other molecules, and the binding can occasionally block and inactivate the bound protein. But antibodies are unable to cross the cell membrane and need an exposed protein outside the cell to bind. Neu, then, was a perfect target, with a large portion, a long molecular "foot," projected tantalizingly outside the cell membrane. It would have taken Padhy no more than an afternoon's experiment to add the neu antibody to the neuroblastoma cells to determine the binding's effect. "It would have been an overnight test," Weinberg would later recall. "I can flagellate myself. If I had been more studious and more focused and not as monomaniacal about the ideas I had at that time, I would have made that connection."
Despite the trail of seductive leads, Padhy and Weinberg never got around to doing their experiment. Afternoon upon afternoon passed. Introspective and bookish, Padhy shuffled through the lab in a threadbare coat in the winter, running his experiments privately and saying little about them to others. And although Padhy's discovery was published in a high-profile scientific journal, few scientists noticed that he might have stumbled on a potential anticancer drug (the neu-binding antibody was buried in an obscure figure in the article). Even Weinberg, caught in the giddy upswirl of new oncogenes and obsessed with the basic biology of the cancer cell, simply forgot about the neu experiment.*
Weinberg had an oncogene and possibly an oncogene-blocking drug, but the twain had never met (in human cells or bodies). In the neuroblastoma cells dividing in his incubators, neu rampaged on monomaniacally, single-mindedly, seemingly invincible. Yet its molecular foot still waved just outside the surface of the plasma membrane, exposed and vulnerable, like Achilles' famous heel.
* In 1986, Jeffrey Drebin and Mark Greene showed that treatment with an anti-neu antibody arrested the growth of cancer cells. But the prospect of developing this antibody into a human anticancer drug eluded all groups.
A City of Strings
In Ersilia, to establish the relationships that sustain the city's life, inhabitants stretch strings from the corners of the houses, white or black or gray or black-and-white according to whether they mark a relationship of blood, of trade, authority, agency. When the strings become so numerous that you can no longer pass among them, the inhabitants leave: the houses are dismantled.
--Italo Calvino
Weinberg may briefly have forgotten about the therapeutic implication of neu, but oncogenes, by their very nature, could not easily be forgotten. In his book Invisible Cities, Italo Calvino describes a fictional metropolis in which every relationship between one household and the next is denoted by a piece of colored string stretched between the two houses. As the metropolis grows, the mesh of strings thickens and the individual houses blur away. In the end, Calvino's city becomes no more than an interwoven network of colored strings.
If someone were to draw a similar map of relationships among genes in a normal human cell, then proto-oncogenes and tumor suppressors such as ras, myc, neu, and Rb would sit at the hub of this cellular city, radiating webs of colored strings in every direction. Proto-oncogenes and tumor suppressors are the molecular pivots of the cell. They are the gatekeepers of cell division, and the division of cells is so central to our physiology that genes and pathways that coordinate this process intersect with nearly every other aspect of our biology. In the laboratory, we call this the six-degrees-of-separation-from-cancer rule: you can ask any biological question, no matter how seemingly distant--what makes the heart fail, or why worms age, or even how birds learn songs--and you will end up, in fewer than six genetic steps, connecting with a proto-oncogene or tumor suppressor.
It should hardly come as a surprise, then, that neu was barely forgotten in Weinberg's laboratory when it was resurrected in another. In the summer of 1984, a team of researchers, collaborating with Weinberg, discovered the human homolog of the neu gene. Noting its resemblance to another growth-modulating gene discovered previously--the Human EGF Receptor (HER)--the researchers called the gene Her-2.
A gene by any other name may still be the same gene, but something crucial had shifted in the story of neu. Weinberg's gene had been discovered in an academic laboratory. Much of Weinberg's attention had been focused on dissecting the molecular mechanism of the neu oncogene. Her-2, in contrast, was discovered on the sprawling campus of the pharmaceutical company Genentech. The difference in venue, and the resulting difference in goals, would radically alter the fate of this gene. For Weinberg, neu had represented a route to understanding the fundamental biology of neuroblastoma. For Genentech, Her-2 represented a route to developing a new drug.
Located on the southern edge of San Francisco, sandwiched among the powerhouse labs of Stanford, UCSF, and Berkeley and the burgeoning start-ups of Silicon Valley, Genentech--short for Genetic Engineering Technology--was born out of an idea imbued with deep alchemic symbolism. In the late 1970s, researchers at Stanford and UCSF had invented a technology termed "recombinant DNA." This technology allowed genes to be manipulated--engineered--in a hitherto unimaginable manner. Genes could be shuttled from one organism to another: a cow gene could be transferred into bacteria, or a human protein synthesized in dog cells. Genes could also be spliced together to create new genes, creating proteins never found in nature. Genentech imagined leveraging this technology of genes to develop a pharmacopoeia of novel drugs. Founded in 1976, the company licensed recombinant DNA technology from UCSF, raised a paltry $200,000 in venture funds, and launched its hunt for these novel drugs.
A "drug," in bare conceptual terms, is any substance that can produce an effect on the physiology of an animal. Drugs can be simple molecules; water and salt, under appropriate circumstances, can function as potent pharmacological agents. Or drugs can be complex, multifaceted chemicals--molecules derived from nature, such as penicillin, or chemicals synthesized artificially, such as aminopterin. Among the most complex drugs in medicine are proteins, molecules synthesized by cells that can exert diverse effects on human physiology. Insulin, made by pancreas cells, is a protein that regulates blood sugar and can be used to control diabetes. Growth hormone, made by the pituitary cells, augments growth by increasing the metabolism of muscle and bone cells.
Before Genentech, protein drugs, although recognizably potent, had been notoriously difficult to produce. Insulin, for instance, was produced by grinding up cow and pig innards into a soup and then extracting the protein from the mix--one pound of insulin from every eight thousand pounds of pancreas. Growth hormone, used to treat a form of dwarfism, was extracted from pituitary glands dissected out of thousands of human cadavers. Clotting drugs to treat bleeding disorders came from liters of human blood.
Recombinant DNA technology allowed Genentech to synthesize human proteins de novo: rather than extracting proteins from animal and human organs, Genentech could "engineer" a human gene into a bacterium, say, and use the bacterial cell as a bioreactor to produce vast quantities of that protein. The technology was transformative. In 1982, Genentech unveiled the first "recombinant" human insulin; in 1984, it produced a clotting factor used to control bleeding in patients with hemophilia; in 1985, it created a recombinant version of human growth hormone--all created by engineering th
e production of human proteins in bacterial or animal cells.
By the late 1980s, though, after an astonishing growth spurt, Genentech ran out of existing drugs to mass-produce using recombinant technology. Its early victories, after all, had been the result of a process and not a product: the company had found a radical new way to produce old medicines. Now, as Genentech set out to invent new drugs from scratch, it was forced to change its winning strategy: it needed to find targets for drugs--proteins in cells that might play a critical role in the physiology of a disease that might, in turn, be turned on or off by other proteins produced using recombinant DNA.
It was under the aegis of this "target discovery" program that Axel Ullrich, a German scientist working at Genentech, rediscovered Weinberg's gene--Her-2/neu, the oncogene tethered to the cell membrane.* But having discovered the gene, Genentech did not know what to do with it. The drugs that Genentech had successfully synthesized thus far were designed to treat human diseases in which a protein or a signal was absent or low--insulin for diabetics, clotting factors for hemophiliacs, growth hormone for dwarfs. An oncogene was the opposite--not a missing signal, but a signal in overabundance. Genentech could fabricate a missing protein in bacterial cells, but it had yet to learn how to inactivate a hyperactive protein in a human cell.
In the summer of 1986, while Genentech was still puzzling over a method to inactivate oncogenes, Ullrich presented a seminar at the University of California in Los Angeles. Flamboyant and exuberant, dressed in a dark, formal suit, Ullrich was a riveting speaker. He floored his audience with the incredible story of the isolation of Her-2, and the serendipitous convergence of that discovery with Weinberg's prior work. But he left his listeners searching for a punch line. Genentech was a drug company. Where was the drug?
Siddhartha Mukherjee - The Emperor of All Maladies: A Biography of Cancer Page 49